Thiophene-carboxamide derivatives and their use as inhibitors of the enzyme IKK-2

ABSTRACT

The invention relates to thiophene carboxamides of formula (I), wherein A, R 1 , R 2 , R 3 , R 4 , R 5  and X are as defined in the specification, processes and intermediates used in their preparation, pharmaceutical compositions containing them and their use in therapy.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is the National Stage of International Application No.PCT/GB2004/000096, filed Jan.13, 2004, which claims the benefit ofSwedish Patent application Ser. No. 0300092-4, filed Jan.15, 2003. Thecontents of both prior applications are hereby incorporated by referencein their entireties.

FIELD OF THE INVENTION

The present invention relates to thiophene carboxamide derivatives,processes and intermediates used in their preparation, pharmaceuticalcompositions containing them and their use in therapy.

BACKGROUND OF THE INVENTION

The NF-κB (nuclear factor κB) family is composed of homo- andheterodimers of the Rel family of transcription factors. A key role ofthese transcription factors is to induce and co-ordinate the expressionof a broad spectrum of pro-inflammatory genes including cytokines,chemokines, interferons, MHC proteins, growth factors and cell adhesionmolecules (for reviews see Verma et. al., Genes Dev. 9:2723-35, 1995;Siebenlist et. al., Ann. Rev. Cell. Biol. 10:405-455, 1994; Bauerle andHenkel, Ann. Rev. Immunol., 12:141-179, 1994; Barnes and Karin, NewEngl. J. Med., 336:1066-1071, 1997).

The most commonly found Rel family dimer complex is composed of p50 NFkBand p65 RelA (Baeuerle and Baltimore, Cell 53:211-217, 1988; Baeuerleand Baltimore, Genes Dev. 3:1689-1698, 1989). Under resting conditionsNF-κB dimers are retained in the cytoplasm by a member of the IκB familyof inhibitory proteins (Beg et. al., Genes Dev., 7:2064-2070, 1993;Gillnore and Morin, Trends Genet. 9:427-433, 1993; Haskil et. al., Cell65:1281-1289, 1991). However, upon cell activation by a variety ofcytokines or other external stimuli, IκB proteins become phosphorylatedon two critical serine residues (Traenckner et. al., EMBO J., 14:2876,1995) and are then targeted for ubiquitination and proteosome-mediateddegradation (Chen, Z. J. et. al., Genes and Dev. 9:1586-1597, 1995;Scherer, D.C. et. al., Proc. Natl. Acad. Sci. USA 92:11259-11263, 1996;Alkalay, I. et. al., Proc. Natl. Acad. Sci. USA 92:10599-10603, 1995).The released NF-κB is then able to translocate to the nucleus andactivate gene transcription (Beg et. al., Genes Dev., 6:1899-1913,1992).

A wide range of external stimuli have been shown to be capable ofactivating NF-κB (Baeuerle, P. A., and Baichwal, V. R., Adv. Immunol.,65:111-136, 1997). Although the majority of NF-κB activators result inIκB phosphorylation, it is clear that multiple pathways lead to this keyevent. Receptor-mediated NF-κB activation relies upon specificinteractions between the receptor and adapter/signaling molecules (forexample, TRADD, RIP, TRAF, MyD88) and associated kinases (IRAK, NIK)(Song et. al., Proc. Natl. Acad. Sci. USA 94:9792-9796, 1997; Natoli et.al., JBC 272:26079-26082, 1997). Environmental stresses such as UV lightand γ-radiation appear to stimulate NF-κB via alternative, less defined,mechanisms.

Recent publications have partially elucidated the NF-κB activation. Thiswork has identified three key enzymes which regulate specific IκB/NF-κBinteractions: NF-κB inducing kinase (NIK) (Boldin et. al., Cell85:803-815, 1996), IκB kinase-1 (IKK-1) (Didonato et. al., Nature388:548, 1997; Regnier at. al., Cell 90:373 1997) and IB kinase-2(IKK-2) (Woronicz et. al., Science 278:866, 1997; Zandi et. al., Cell91:243, 1997).

NIK appears to represent a common mediator of NF-κB signaling cascadestriggered by tumour necrosis factor and interleukin-1, and is a potentinducer of IκB phosphorylation. However NIK is unable to phosphorylateIκB directly.

IKK-1 and IKK-2 are thought to lie immediately downstream of NIK and arecapable of directly phosphorylating all three IκB sub-types. IKK-1 andIKK-2 are 52% identical at the amino acid level but appear to havesimilar substrate specificities; however, enzyme activities appear to bedifferent: IKK-2 is several-fold more potent than IKK-1. Expressiondata, coupled with mutagenesis studies, suggest that IKK-1 and IKK-2 arecapable of forming homo- and heterodimers through their C-terminalleucine zipper motifs, with the heterodimeric form being preferred(Mercurio et. al., Mol. Cell Biol., 19:1526, 1999; Zandi et. al.,Science; 281:1360, 1998; Lee et. al, Proc. Natl. Acad. Sci. USA 95:9319,1998).

NIK, IKK-1 and IKK-2 are all serine/threonine kinases. Recent data hasshown that tyrosine kinases also play a role in regulating theactivation of NF-κB. A number of groups have shown that TNF-α inducedNF-κB activation can be regulated by protein tyrosine phosphatases(PTPs) and tyrosine kinases (Amer et. al., JBC 273:29417-29423, 1998; Huet. al., JBC 273:33561-33565, 1998; Kaekawa et. al., Biochem. J.337:179-184, 1999; Singh et. al., JBC 271 31049-31054, 1996). Themechanism of action of these enzymes appears to be in regulating thephosphorylation status of IκB. For example, PTP1B and an unidentifiedtyrosine kinase appear to directly control the phosphorylation of alysine residue (K42) on IκB-α, which in turn has a critical influence onthe accessibility of the adjacent serine residues as targets forphosphorylation by IKK.

Several groups have shown that IKK-1 and IKK-2 form part of a‘signalosome’ structure in association with additional proteinsincluding IKAP (Cohen et. al., Nature 395:292-296, 1998; Rothwarf et.al., Nature 395:297-300, 1998), MEKK-1, putative MAP kinase phosphatase(Lee et. al., Proc. Natl. Acad. Sci. USA 95:9319-9324, 1998), as well asNIK and IκB. Data is now emerging to suggest that although both IKK-1and IKK-2 associate with NIK, they are differentially activated, andtherefore might represent an important integration point for thespectrum of signals that activate NF-κB. Importantly, MEKK-1 (one of thecomponents of the putative signalosome and a target for UV light, LPSinduced signaling molecules and small GTases) has been found to activateIK-2 but not IKK-1. Similarly, NIK phosphorylation of IKK-1 results in adramatic increase in IKK-1 activity but only a small effect on IKK-2(for review, see Mercurio, F., and Manning, A. M., Current Opinion inCell Biology, 11:226-232, 1999).

Inhibition of NF-κB activation is likely to be of broad utility in thetreatment of inflammatory disease.

There is accumulating evidence that NF-κB signaling plays a significantrole in the development of cancer and metastasis. Abnormal expression ofc-Rel, NF-κB2 or IκBα have been described in a number of tumour typesand tumour cell lines, and there is now data to show that constitutiveNF-κB via IKK-2 takes place in a wide range of tumour cell lines. Thisactivity has been linked to various upstream defects in growth factorsignaling such as the establishment of autocrine loops, or the presenceof oncogene products e.g. Ras, AKT, Her2, which are involved in theactivation of the IKK complex. Constitutive NF-κB activity is believedto contribute to oncogenesis through activation of a range ofanti-apoptotic genes e.g. A1/Bfi-1, IEX-1, XIAP, leading to thesuppression of cell death pathways, and transcriptional upregulation ofcyclin D1 which promotes cell growth. Other data indicate that thispathway is also likely to be involved in the regulation of cell adhesionand cell surface proteases. This suggests a possible additional role forNF-κB activity in the development of metastasis. Evidence confirming theinvolvement of NF-κB activity in oncogenesis includes the inhibition oftumour cell growth in vitro and in vivo on expression of a modified formof IκBα (super-repressor IκBα).

In addition to the constitutive NF-κB signaling observed in many tumourtypes, it has been reported that NF-κB is also activated in response tocertain types of chemotherapy. Inhibition of NF-κB activation throughexpression of the super-repressor form of IκBα in parallel withchemotherapy treatment has been shown to enhance the anti-tumour effectof the chemotherapy in xenograft models. NF-κB activity is thereforealso implicated in inducible chemoresistance.

Patent application WO 01/58890 discloses certain thiophene carboxamidederivatives that are useful as IKK-2 inhibitors. We now disclose afurther group of thiophene carboxamide derivatives that possessdesirable pharmacological activity profiles, in particular, increasedbeneficial potencies.

DISCLOSURE OF THE INVENTION

According to the present invention, there is provided a compound offormula (I)

wherein

-   R¹ represents H or CH₃;-   R² represents hydrogen, halogen, cyano, C1 to 2 alkyl,    trifluoromethyl or C1 to 2 alkoxy;-   R³ and R⁴ independently represent H or CH₃; or the group CR³R⁴    together represents a C3 to 6 cycloalkyl ring;-   A represents a six-membered aromatic ring optionally incorporating    one or two nitrogen atoms; and the group —CR³R⁴—X—R⁵ is bonded to    ring A in the 4-position relative to the thiophene ring;-   X represents NR⁶;-   R⁵ represents H, C1 to 6 alkyl, C2 to 6 alkenyl or C3 to 6    cycloalkyl; said cycloalkyl group optionally incorporating one    heteroatom selected from O, S(O)_(n) or NR⁷; said alkyl group being    optionally further substituted by one or more groups selected    independently from CN, OH, C1 to 4 alkoxy, F, a C5 to 10 monocyclic    or bicyclic aromatic ring system optionally incorporating one or two    heteroatoms independently selected from O, S and N, and said ring    system being optionally further substituted by one or more    substituents selected independently from halogen, C1 to 2 alkyl, C1    to 2 alkoxy or CF₃; or said alkyl being optionally further    substituted by a C5 to 6 cycloalkyl ring that optionally    incorporates a heteroatom selected from O, S(O)_(m) or NR⁸ and/or a    carbonyl group and is optionally further substituted by OH; R⁶    represents H or C1 to 6 alkyl; said alkyl group being optionally    further substituted by CN, OH, CI to 4 alkoxy or one or more fluoro    atoms;-   n and m independently represent an integer 0, 1 or 2;-   R⁷ and R⁸ independently represent H or C1 to 2 alkyl;    and pharmaceutically acceptable salts thereof.

Certain compounds of formula (I) are capable of existing instereoisomeric forms. It will be understood that the inventionencompasses all geometric and optical isomers of the compounds offormula (I) and mixtures thereof including racemates. Tautomers andmixtures thereof also form an aspect of the present invention.

In one embodiment, R¹ in formula (I) represents H.

In another embodiment, R² in formula (I) represents H.

In another embodiment, A represents an optionally substituted phenyl.

In another embodiment, R³ and R⁴ in formula (I) each represent H.

In yet another embodiment, R⁵ represents C1 to 4 alkyl or C3 to 6cycloalkyl; said cycloalkyl group optionally incorporating oneheteroatom selected from O, S(O)_(n) or NR⁷; and said alkyl group beingoptionally further substituted by 1, 2 or 3 groups selectedindependently from CN, OH, C1 to 4 alkoxy, F, a C5 to 10 monocyclic andbicyclic aromatic ring system optionally incorporating one or twoheteroatoms independently selected from O, S and N, and said ring systembeing optionally further substituted by 1, 2 or 3 substituents selectedindependently from halogen, C1 to 2 alkyl C1 to 2 alkoxy and CF₃; orsaid alkyl being optionally further substituted by a C5 to 6 cycloalkylring that optionally incorporates a heteroatom selected from O, S(O)_(m)or NR⁸ and/or a carbonyl group and is optionally further substituted by1 OH group.

In another embodiment, R⁵ represents C1 to 6 alkyl or C3 to 6cycloalkyl; said cycloalkyl group optionally incorporating oneheteroatom selected from O, S(O)_(n) or NR⁷; and said alkyl group beingoptionally further substituted by 1, 2 or 3 groups selectedindependently from CN, OH, C1 to 4 alkoxy, F, a C5 to 10 monocyclic anda bicyclic aromatic ring system optionally incorporating one or twoheteroatoms independently selected from O, S and N, and said ring systembeing optionally further substituted by 1, 2 or 3 substituents selectedindependently from halogen, C1 to 2 alkyl, C1 to 2 alkoxy and CF₃; orsaid alkyl being optionally further substituted by a C5 to 6 cycloalkylring that optionally incorporates a heteroatom selected from O, S(O)_(m)or NR⁸ and/or a carbonyl group and is optionally further substituted by1 OH group.

In another embodiment R⁶ represents H or C1 to 4 alkyl; said alkyl groupbeing optionally substituted by CN, OH, C1 to 4 alkoxy or by 1, 2 or 3fluoro atoms.

In one embodiment, the carboxamido group in formula (I) is attached tothe 3-position of the thiophene ring.

In another embodiment, the carboxamido group in formula (I) is attachedto the 2-position of the thiophene ring.

In one embodiment the present invention relates to a class of compoundsof formula (I) wherein R¹, R², R³ and R⁴ each represent H; thecarboxamido group is attached to the 3-position of the thiophene ring;the carboxamido group in formula (I) is attached to the 2-position ofthe thiophene ring; A represents an optionally substituted phenyl; andR⁵ and R⁶ are as defined above.

In another embodiment the present invention relates to a class ofcompounds of formula (I) wherein R¹, R², R³ and R⁴ each represent H; thecarboxamido group is attached to the 3-position of the thiophene ring;the carboxamido group in formula (I) is attached to the 2-position ofthe thiophene ring; A represents phenyl; R6 is H or C1 to 4 alkyl; saidalkyl group being optionally substituted by CN, OH, C1 to 4 alkoxy or by1, 2 or 3 fluoro atoms; and R⁵ has any of the definitions defined above.

In another embodiment the present invention relates to a class ofcompounds of formula (I) wherein R¹, R², R³ and R⁴ each represent H; thecarboxamido group is attached to the 3-position of the thiophene ring;the carboxamido group in formula (I) is attached to the 2-position ofthe thiophene ring; A represents phenyl; R6 is H or C1 to 4 alkyl; saidalkyl group being optionally substituted by CN, OH, C1 to 4 alkoxy or by1, 2 or 3 fluoro atoms; and R⁵ represents C1 to 6 alkyl or C3 to 6cycloalkyl; said cycloalkyl group optionally incorporating oneheteroatom selected from O, S(O)_(n) or NR⁷; and said alkyl group beingoptionally further substituted by 1, 2 or 3 groups selectedindependently from CN, OH, C1 to 4 alkoxy, F, a C5 to 10 monocyclic anda bicyclic aromatic ring system optionally incorporating one or twoheteroatoms independently selected from O, S and N, and said ring systembeing optionally further substituted by 1, 2 or 3 substituents selectedindependently from halogen, C1 to 2 alkyl, C1 to 2 alkoxy and CF₃; orsaid alkyl being optionally further substituted by a C5 to 6 cycloalkylring that optionally incorporates a heteroatom selected from O, S(O)_(m)or NR⁸ and/or a carbonyl group and is optionally further substituted by1 OH group.

The compounds of formula (I) and their pharmaceutically acceptable saltshave the advantage that they are inhibitors of the enzyme IKK-2.

The invention further provides a process for the preparation ofcompounds of formula (I) or a pharmaceutically acceptable salt,enantiomer or racemate thereof.

According to the invention there is also provided a compound of formula(I), or a pharmaceutically acceptable salt thereof for use as amedicament.

Another aspect of the invention provides the use of a compound offormula (I) or a pharmaceutically acceptable salt thereof, in themanufacture of a medicament, for the treatment or prophylaxis ofdiseases or conditions in which inhibition of IKK-2 activity isbeneficial.

A more particular aspect of the invention provides the use of a compoundof formula (I) or a pharmaceutically acceptable salt thereof, in themanufacture of a medicament, for the treatment or prophylaxis ofinflammatory disease.

According to the invention, there is also provided a method of treating,or reducing the risk of, diseases or conditions in which inhibition ofIKK-2 activity is beneficial which comprises administering to a personsuffering from or at risk of, said disease or condition, atherapeutically effective amount of a compound of formula (I), or apharmaceutically acceptable salt thereof.

More particularly, there is also provided a method of treating, orreducing the risk of inflammatory disease in a person suffering from orat risk of, said disease, wherein the method comprises administering tothe person a therapeutically effective amount of a compound of formula(I) or a pharmaceutically acceptable salt thereof.

Particular compounds of the invention include those exemplified herein:

-   2-[(aminocarbonyl)amino]-5-(4-{[2,2,2-trifluoroethyl)amino]methyl}phenyl)thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-{4-[(isopropylamino)methyl]phenyl}thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-{4-[(bis-(2-methoxyethyl)amino)methyl]phenyl}thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-{4-[(N-ethyl-N-(2-methoxyethyl)amino)methyl]phenyl}-thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-{4-[dimethylaminomethyl]phenyl}thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-{4-[(N-(2,2,2-trifluoroethyl)-N-(2-methoxyethyl)amino)methyl]phenyl}thiophene-3    -carboxamide;-   2-[(aminocarbonyl)amino]-5-{4-[(2-methoxyethyl)amino)methyl]phenyl}thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(2-(1H-indol-3-yl)ethyl)amino]methyl}phenyl)    thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{methylaminomethyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{cyclopropylaminomethyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{([(2R)-2-hydroxypropyl]amino)methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{([(2S)-2-hydroxypropyl]amino)methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(tetrahydrofuran-2-ylmethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(4-fluorobenzyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(3-{2-oxopyrrolidin-1-yl}propyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(1-naphthylmethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{([1-(4-chlorobenzyl)-2-hydroxyethyl]amino)methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{(cyclopentylamino)methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(pyridin-3-ylmethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(2-[pyridin-2-yl]ethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[({2-hydroxy-1,1-dimethyl}ethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[({1,2-diphenyl}ethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[({2-methoxy-1-methyl}ethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[({2-hydroxy-1-methyl}ethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(2-methylbenzyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(3-methoxybenzyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(2-fluorobenzyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(3-fluorobenzyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(4-{phenyl}butyl)amino]methyl}phenyl)thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{([3-(trifluoromethyl)benzyl]amino)methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(5-cyanopentyl)amino)methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{({2-methyl}propylamino)methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(4-methoxybenzyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(2-phenylethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[(2-hydroxyethyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[({2-methoxy-2-methyl}propyl)amino]methyl}phenyl]thiophene-3-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-(4-fluorobenzyl)-N-methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-(2-{pyridin-2-yl)    ethyl)methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-(pyridin-2-ylmethyl)methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-benzyl-N-methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-(2-methoxyethyl)ethylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[(N-{2-methoxyethyl})methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-benzyl-N-(2-cyanoethyl)amino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-(2-cyanoethyl)-N-methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-isopropyl-N-(2-methoxyethyl)amino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[(3-methoxybenzyl)amino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-(2-methoxyethyl)-N-(pyridin-2-yl)methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-(3,5-dimethyl-1H-pyrazol-yl)methyl]methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{({N-[3-methylisoxazol-5-yl]methyl}methylamino)    methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-(2-hydroxyethyl)methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[(2-methoxyethyl)amino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-(1,1-dioxotetrahydro-3-thienyl)methylamino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[N-benzyl-N-(2-hydroxyethyl)amino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[(tetrahydrofuran-2-ylmethyl)amino]methyl}phenyl]thiophene-2-carboxamide;-   3-[(aminocarbonyl)amino]-5-[4-{[(2-methoxy-2-methyl)propylamino]methyl}phenyl]thiophene-2-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[N-(2-methoxyethyl)methylamino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{(N,N-diethylamino)methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{([N-(2-cyanoethyl)-N-methylamino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[N-benzyl-N-cyanoethylamino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{[N-(2-hydroxyethyl)-N-methylamino]methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{([N-benzyl-N-(2-hydroxyethyl)]amino)methyl}phenyl]thiophene-3-carboxamide;-   2-[(aminocarbonyl)amino]-5-[4-{(bis-[2-hydroxyethyl]amino)methyl}phenyl]thiophene-3-carboxamide;    and pharmaceutically acceptable salts thereof.

Unless otherwise indicated, the term “C1 to 6 alkyl” referred to hereindenotes a straight or branched chain alkyl group having from 1 to 6carbon atoms. Examples of such groups include methyl, ethyl, n-propyl,i-propyl, n-butyl, i-butyl and t-butyl. The term “C1 to 2 alkyl” is tobe interpreted analogously.

Unless otherwise indicated, the term “C2 to 6 alkenyl” referred toherein denotes a straight or branched chain alkyl group having 2 to 6carbon atoms incorporating at least one carbon-carbon double bond.Examples of such groups include ethenyl and propenyl.

Unless otherwise indicated, the term “C3 to 6 cycloalkyl” referred toherein denotes a 5 saturated carbocyclic ring having from 3 to 6 carbonatoms. Examples of such groups include cyclopropyl, cyclopentyl andcyclohexyl.

Unless otherwise indicated, the term “C1 to 4 alkoxy” referred to hereindenotes a straight or branched chain alkoxy group having 1 to 4 carbonatoms. Examples of such groups include methoxy, ethoxy and isopropoxy.The term “C1 to 2 alkoxy” is to be interpreted analogously.

Unless otherwise indicated, the term “halogen” referred to hereindenotes fluoro, chloro, bromo and iodo.

Examples of a six-membered aromatic ring optionally incorporating one ortwo nitrogen atoms include phenyl, pyridine, pyridazine, pyrimidine andpyrazine.

Examples of a C3 to 6 cycloalkyl ring that optionally incorporates oneheteroatom selected from O, S(O)_(n) or NR⁸ include cyclopropyl,cyclopentyl, cyclohexyl, tetrahydrofuranyl, tetrahydropyranyl,tetrahydrothienyl-1,1-dioxide, pyrrolidinyl, pyrrolidonyl andpiperidinyl.

Examples of a C5 to 10 monocyclic or bicyclic aromatic ring systemoptionally incorporating one or two heteroatoms independently selectedfrom O, S and N include phenyl, pyridyl, naphthyl, indolyl, isoxazolyland pyrazolyl Examples of a C5 to 6 cycloalkyl ring that optionallyincorporates a heteroatom selected from O, S(O)_(m) or NR⁸ and/or acarbonyl group include cyclohexyl, tetrahydrofuranyl, tetrahydropyranyl,tetrahydrothienyl 1,1-dioxide, pyrrolidinyl, pyrrolidonyl andpiperidinyl.

According to the invention there is also provided a process for thepreparation of a compound of formula (I) or a pharmaceuticallyacceptable salt, enantiomer or racemate thereof which comprises:

-   (a) reaction of a compound of formula (II):

wherein A, R¹, R², R³, R⁴, R⁵ and X are as defined in formula (I) withan isocyanate; or

-   (b) reaction of compound of formula (III)

wherein A, R², R³, R⁴, R⁵ and X are as defined in formula (I), with acompound of formula (IV)

wherein R¹ is as defined in formula (I) and LG represents a leavinggroup; or

-   (c) reaction of compound of formula (V)

wherein A, R², R³, R⁴, R⁵ and X are as defined in formula (I) and LGrepresents a leaving group,with a compound of formula (VI)

wherein R¹ is as defined in formula (I); or

-   (d) reaction of compound of formula (VII)

wherein A, R¹, R², R³ and R⁴ are as defined in formula (I), and LGrepresents a leaving group, with an amine of formula R⁵R⁶NH, wherein R⁵and R⁶ are as defined in formula (I); or

-   (e) reaction of compound of formula (VII)

wherein A, R¹, R² and R³ are as defined in formula (I),with an amine of formula R⁵R⁶NH wherein R⁵ and R⁶ are as defined informula (I), under reductive amination conditions; or

-   (f) reaction of a compound of formula (IX)

wherein R¹, R², R³, R⁴, R⁵ and A are as defined in formula (I), with analdehyde or ketone under reductive amination conditions;

-   -   and where necessary converting the resultant compound of formula        (I), or another salt thereof into a pharmaceutically acceptable        salt thereof; or converting the resultant compound of        formula (I) into a further compound of formula (I); and where        desired converting the resultant compound of formula (I) into an        optical isomer thereof.

In process (a), suitable isocyanate reagents includetrimethylsilylisocyanate, trichloroacetylisocyanate and sodiumisocyanate. The reaction with trimethylsilylisocyanate can be carriedout in a solvent such as dichloromethane/dimethylformamide at a suitableelevated temperature, for example, at the reflux temperature of thereaction mixture. The reaction with sodium isocyanate can be carried outin a suitable solvent system such as aqueous acetic acid at ambienttemperature. The trichloroacetylisocyanate reaction can be carried outin a suitable solvent system such as acetonitrile at ambienttemperature, and subsequently treating the mixture with ammonia to givecompounds of the general formula (I). In a preferred embodiment, theisocyanate is trichloroacetylisocyanate.

In processes (b) and (c), the compounds of formulae (II) and (V) or offormulae (V) and (VI) are reacted together under catalysis provided by acomplex of a transition metal such as palladium or nickel. In compoundsof formulae (III) and (VI), under appropriate conditions, “metal” can bea metal or semi-metal such as magnesium, zinc, copper, tin, silicon,zirconium, aluminium or boron. Suitable leaving groups include iodo,bromo, chloro, triflate or phosphonate.

In process (d), the compounds of formulae (VII) are reacted togetherwith amines under appropriate reaction conditions. This can either be inthe presence or absence of base. Such bases can be either inorganic ororganic. Suitable leaving groups include iodo, bromo, chloro, sulphonateand triflate.

In process (e), the carbonyl compounds of formula (VIM) are reactedtogether with amines under appropriate reductive amination reactionconditions. The reducing agent for these reactions include sodiumcyanoborohydride and sodium triacetoxyborohydride. Solvents can includetrimethylorthoformate and methanol. Titanium (IV) salts may also be usedin this process. Alternatively, compounds of the formula (VIII) can bereacted with amines to form the corresponding imine, which can then bereduced to produce compounds of formula (I). For this route, additionalreducing agents such as sodium borohydride may be used.

Conditions for process (f) are analogous to those described above forprocess (e).

It will be appreciated by those skilled in the art that in the processesof the present invention certain functional groups such as hydroxyl oramino groups in the starting reagents or intermediate compounds may needto be protected by protecting groups. Thus, the preparation of thecompounds of formula (I) may involve, at an appropriate stage, theaddition and removal of one or more protecting groups.

The protection and deprotection of functional groups is described in‘Protective Groups in Organic Chemistry’, edited by J. W. F. McOmie,Plenum Press (1973), and ‘Protective Groups in Organic Synthesis’, 3rdedition, T. W. Greene & P. G. M. Wuts, Wiley-Interscience (1999).

The present invention includes compounds of formula (I) in the form ofsalts, in particular acid addition salts. Suitable salts include thoseformed with both organic and inorganic acids. Such acid addition saltswill normally be pharmaceutically acceptable although salts ofnon-pharmaceutically acceptable acids may be of utility in thepreparation and purification of the compound in question. Thus,preferred salts include those formed from hydrochloric, hydrobromic,sulphuric, phosphoric, citric, tartaric, lactic, pyruvic, acetic,succinic, fumaric, maleic, methanesulphonic and benzenesulphonic acids.

Salts of compounds of formula (I) may be formed by reacting the freebase, or a salt, enantiomer or racemate thereof, with one or moreequivalents of the appropriate acid. The reaction may be carried out ina solvent or medium in which the salt is insoluble or in a solvent inwhich the salt is soluble, for example, water, dioxane, ethanol,tetrahydrofuran or diethyl ether, or a mixture of solvents, which may beremoved in vacuo or by freeze drying. The reaction may also be ametathetical process or it may be carried out on an ion exchange resin.

Compounds of formula (II) can be prepared by standard chemistrydescribed in the literature [for example, J. Heterocyclic Chem., 36, 333(1999)] or by reaction of compounds of formula (X)

where A, R¹, R², R³, R⁴, R⁵ and X are as defined in formula (I) with ahalogenating agent such as thionyl chloride.

Compounds of formulae (III), (T), (V), (VI), (VII), (VIII) and (IX) arecommercially available or can be prepared using standard chemistry asexemplified herein.

Certain novel intermediate compounds form a further aspect of theinvention.

The compounds of formula (I) have activity as pharmaceuticals, inparticular as IKK-2 enzyme inhibitors, and may be used in the treatment(therapeutic or prophylactic) of conditions/diseases in human andnon-human animals in which inhibition of IKK-2 is beneficial. Examplesof such conditions/diseases include inflammatory diseases or diseaseswith an inflammatory component. Particular diseases include inflammatoryarthritides including rheumatoid arthritis, osteoarthritis, spondylitis,Reiters syndrome, psoriatic arthritis, lupus and bone resorptivedisease; multiple sclerosis, inflammatory bowel disease includingCrohn's disease; asthma, chronic obstructive pulmonary disease,emphysema, rhinitis, myasthenia gravis, Graves' disease, allograftrejection, psoriasis, dermatitis, allergic disorders, immune complexdiseases, cachexia, ARDS, toxic shock, heart failure, myocardialinfarcts, atherosclerosis, reperfusion injury, AIDS, cancer anddisorders characterised by insulin resistance such as diabetes,hyperglycemia, hyperinsulinemia, dyslipidemia, obesity, polycysticovarian disease, hypertension, cardiovascular disease and Syndrome X.

The reported roles of NF-κB in both oncogenesis and chemoresistancesuggest that inhibition of this pathway through the use of an IKK-2inhibitor, such as a small molecule IKK-2 inhibitor, could provide anovel monotherapy for cancer and/or an important adjuvant therapy forthe treatment of chemoresistant tumours and in the synergistic inductionof apoptosis as a result of combination therapy with an IKK-2 inhibitorwith standard therapies or other novel agents.

We are particularly interested in diseases selected from asthma,rheumatoid arthritis, psoriasis, inflammatory bowel disease includingCrohn's disease, multiple sclerosis, chronic obstructive pulmonarydisease, bone resorptive disease, osteoarthritis, diabetes/glycaemiccontrol and cancer.

Thus, the present invention provides a compound of formula (I), or apharmaceutically acceptable salt thereof, as hereinbefore defined foruse in therapy.

In a further aspect, the present invention provides the use of acompound of formula (I), or a pharmaceutically acceptable salt thereof,as hereinbefore defined in the manufacture of a medicament for use intherapy.

In a still further aspect, the present invention provides the use of acompound of formula (I), or a pharmaceutically acceptable salt thereof,as hereinbefore defined in the manufacture of a medicament for thetreatment of diseases or conditions in which modulation of the IKK-2enzyme activity is beneficial.

In the context of the present specification, the term “therapy” alsoincludes “prophylaxis” unless there are specific indications to thecontrary. The terms “therapeutic” and “therapeutically” should beconstrued accordingly.

Prophylaxis is expected to be particularly relevant to the treatment ofpersons who have suffered a previous episode of, or are otherwiseconsidered to be at increased risk of, the disease or condition inquestion. Persons at risk of developing a particular disease orcondition generally include those having a family history of the diseaseor condition, or those who have been identified by genetic testing orscreening to be particularly susceptible to developing the disease orcondition.

The invention still further provides a method of treating an IKK-2mediated disease which comprises administering to a patient atherapeutically effective amount of a compound of formula (I), or apharmaceutically acceptable salt thereof, as hereinbefore defined.

The invention also provides a method of treating an inflammatorydisease, especially asthma, rheumatoid arthritis or multiple sclerosis,in a patient suffering from, or at risk of, said disease, whichcomprises administering to the patient a therapeutically effectiveamount of a compound of formula (I), or a pharmaceutically acceptablesalt thereof, as hereinbefore defined

For the above-mentioned therapeutic uses the dosage administered will,of course, vary with the compound employed, the mode of administration,the treatment desired and the disorder indicated.

The compounds of formula (I) and pharmaceutically acceptable saltsthereof may be used on their own but will generally be administered inthe form of a pharmaceutical composition in which the formula (I)compound/salt (active ingredient) is in association with apharmaceutically acceptable adjuvant, diluent or carrier. Depending onthe mode of administration, the pharmaceutical composition willpreferably comprise from 0.05 to 99% w (percent by weight), morepreferably from 0.05 to 80% w, still more preferably from 0.10 to 70% w,and even more preferably from 0.10 to 50% w, of active ingredient, allpercentages by weight being based on total composition.

The present invention also provides a pharmaceutical compositioncomprising a compound of formula (I), or a pharmaceutically acceptablesalt thereof, as hereinbefore defined, in association with apharmaceutically acceptable adjuvant, diluent or carrier.

The invention further provides a process for the preparation of apharmaceutical composition of the invention, which comprises mixing acompound of formula (I), or a pharmaceutically acceptable salt thereof,as hereinbefore defined, with a pharmaceutically acceptable adjuvant,diluent or carrier.

The pharmaceutical compositions may be administered topically (e.g. tothe lung and/or airways or to the skin) in the form of solutions,suspensions, heptafluoroalkane aerosols and dry powder formulations; orsystemically, e.g. by oral administration in the form of tablets,capsules, syrups, powders or granules, or by parenteral administrationin the form of solutions or suspensions, or by subcutaneousadministration or by rectal administration in the form of suppositoriesor transdermally. Conventional procedures for the selection andpreparation of suitable pharmaceutical formulations are described in,for example, “Pharmaceuticals—The Science of Dosage Form Designs”, M. E.Aulton, Churchill Livingstone, 1988.

In one aspect of the invention the composition may be adapted foradministration by inhalation or insufflation. For example, thecomposition may be administered in a form suitable for inhalation, forexample as a finely divided powder or a liquid aerosol such as anaerosol formed from a predominantly aqueous solution or suspension, orfor administration by insufflation, for example as a finely dividedpowder.

It will be appreciated that delivery by inhalation or insufflationprovides higher concentrations of the drug to the required site, namelythe epithelial lining of the lungs, than those readily achievablefollowing systemic absorption of the drug. Smaller doses can thereforebe used to delivered the drug locally to the specific cells which are tobe controlled. Thereby, any adverse systemic side effects of the drugare reduced and the beneficial effects of the treatment can be realisedmore quickly.

Such administration may use a compressed gas to expel the drug from acontainer, for example an aerosol formulation may be used comprisingfine liquid or solid particles carried by a propellant gas underpressure. The aerosol contains the drug which is dissolved, suspended oremulsified in a mixture of a fluid carrier and a propellant.Conventional propellants may be used, for example hydrocarbons or othersuitable gases or mixtures thereof. Conventional metered dose aerosoland breath-activated delivery devices (MDIs) may be employed.Alternatively, the drug may be administered using a conventionalnebuliser, which generates fine liquid particles of substantiallyuniform size containing the drug dispersed as small droplets that canpenetrate into the respiratory tract of the patient.

Alternatively, a powder composition containing the drug, with or withouta lubricant, carrier or propellant, may be used. For example, a powdermixture of the compound and a suitable powder base such as lactose orstarch may be presented in a unit dosage form that may be administeredwith the aid of an inhaler.

However, certain patients may produce copious quantities of mucus in thelungs and such patients may not be treatable initially by inhalation. Inthat event, it may be preferable to delivery the pharmaceuticalcomposition of the present invention by injection or orally.

The compositions of the invention may be obtained by conventionalprocedures using conventional pharmaceutical excipients that are wellknown in the art. Thus, compositions intended for oral use may contain,for example, one or more colouring, sweetening, flavouring and/orpreservative agents.

The invention is illustrated by the following examples: The followingabbreviations are used.

-   DCM Dichloromethane;-   DMA N,N-dimethylacetamide;-   DME 1,2-Dimethoxyethane;-   DMF N,N-Dimethylformamide;-   THF Tetrahydrofuran.

Unless otherwise indicated, organic solutions were dried using anhydrousmagnesium sulphate.

EXAMPLE 12-[(Aminocarbonyl)amino]-5-(4-{[2,2,2-trifluoroethyl)amino]methyl}phenyl)thiophene-3-carboxamide

a) N-(4-Bromobenzyl)-N-(2,2,2-trifluoroethyl)amine

4-Bromobenzylbromide (1.5 g) was stirred with 2,2,2-trifluoroethylamine(0.48 ml) and 10 potassium carbonate (0.99 g) in DMA (5 ml) at roomtemperature for 18 h. The reaction mixture was then poured into water(40 ml) and extracted with ether (3×20 ml). The combined extracts werewashed with water, dried, filtered and evaporated. The residue waspurified by column chromatography, eluting with a gradient of 3 to 5%ethyl acetate in hexane, to afford the product as a viscous, colourlessoil (0.885 g).MS (ES) 268 (M+H)⁺.¹H NMR (CDCl₃) 3.15 (q, 2H), 3.87 (s, 2H), 7.20 (d, 2H), 7.46 (d, 2H).b)2-[(Aminocarbonyl)amino]-5-(4-{[2,2,2-trifluoroethyl)amino]methyl}phenyl)thiophene-3-carboxamideA mixture of N-(4-bromobenzyl)-N-(2,2,2-trifluoroethyl)amine (0.885 g),bis-(pinacolato)diboron (1.68 g), potassium acetate (0.97 g) andPdCl₂(dppf) (0.097 g) in DMA (15 ml) was flushed with argon, heated at80° C. for 4 h and then allowed to cool.2-[(Aminocarbonyl)amino]-5-bromo-3-thiophenecarboxamide (0.349 g) wasadded, followed by a further portion of PdCl₂(dppf) (0.097 g) and 2Maqueous sodium hydrogen carbonate (4 ml). The mixture was heated at 90°C. for 18 h and allowed to cool to room temperature. The solvent wasremoved in vacuo and the residue taken up in 2M aqueous sodium hydroxide(50 ml) and DCM (40 ml). The layers were separated and the aqueous layerwas washed with further DCM (20 ml). The aqueous phase was filtered toremove a small amount of insoluble material and the filtrate thenneutralised with concentrated hydrochloric acid. The precipitatedproduct was collected by filtration and washed with water and dried. Thecrude material was purified by preparative hplc, product-containingfractions combined, neutralised with concentrated aqueous ammonia andevaporated to give the product as a pale brown solid (0.008 g).MS (ES) 371 (M−H)⁻.¹H NMR (DMSO-D6) 2.84-2.95 (m, 1H), 3.11-3.30 (m, 2H), 3.79 (d, 2H),6.98 (bs, 2H), 7.30 (bs, 1H), 7.36 (d, 2H), 7.50 (d, 2H), 7.67 (bs, 1H),7.70 (s, 1H), 11.00 (s, 1H).

EXAMPLE 22-[(Aminocarbonyl)amino]-5-{4-[(isopropylamino)methyl]phenyl}thiophene-3-carboxamide

a)N-[4-(4,4,5,5-Tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl]propan-2-amine

Isopropylamine (0.5 ml) was added dropwise to a solution of2-[4-(bromomethyl)phenyl]-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (0.5g) in dimethoxymethane (10 ml) and the resulting solution was stirred atroom temperature for 18 h. The reaction mixture was evaporated underreduced pressure and used immediately.b)2-[(Aminocarbonyl)amino]-5-{4-[(isopropylamino)methyl]phenyl}thiophene-3-carboxamideA solution ofN-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl]propan-2-amine(0.46 g), 2-[(aminocarbonyl)amino]-5-bromothiophene-3-carboxamide (0.2g) and sodium carbonate (0.32 g) in a DME (15 ml)/water (1.5 ml) mixturewas heated to 80° C. under an argon stream.Tetrakis-(triphenylphosphine)palladium(0) (5.1 g) was then added and thereaction was stirred at 90° C. for 4 h, cooled and evaporated underreduced pressure. The residue was partitioned between DCM and saturatedsodium carbonate and the solid interlayer was filtered and washed withwater. The pure product was obtained by cation exchange chromatographyeluting with ammonia/methanol mixtures (0.06 g).MS (ES) 333 (M+H)⁺.¹H NMR (DMSO-D6) 1.0 (d, 6H), 2.7 (m, 1H), 3.7 (s, 2H), 6.9 (br, 2H),7.3 (br, 1H), 7.4 (d, 2H), 7.5 (d, 2H), 7.7 (br, 1H), 7.75 (s, 1H), 11.0(s, 1H).

EXAMPLE 32-[(Aminocarbonyl)amino]-5-{4-[(bis-(2-methoxyethyl)amino)methyl]phenyl}thiophene-3-carboxamide

a) N-(4-Bromobenzyl)-2-methoxy-N-(2-methoxyethyl)ethanamide

Di-(2-methoxyethyl)amine (6 ml) was added to a solution of4-bromobenzylbromide (5 g) in DMF (20 ml) at ambient temperature. Afterstirring at ambient temperature for 2 h, the mixture was partitionedbetween ethyl acetate and saturated sodium hydrogen carbonate solution.The organic phase was dried, the solvent removed under vacuum to yield ayellow oil which was purified by column chromatography using a 0 to 20%ethyl acetate/isohexane gradient to yield the title compound as a clearoil (4.83 g). MS (M+H)⁺302.¹H NMR (CDCl₃) 2.72 (t, 4H), 3.3 (s, 6H), 3.45 (t, 4H), 3.64 (s, 2H),7.21 (d, 2H), 7.41 (d, 2H).b)2-[(Aminocarbonyl)amino]-5-{4-[(bis-(2-methoxyethyl)amino)methyl]phenyl}-thiophene-3-carboxamideA solution of N-(4-bromobenzyl)-2-methoxy-N-(2-methoxyethyl)ethanamide(1.6 g) in THF (15 ml) was cooled to −60° C. under argon. Butyl lithium(5 ml, 1.6M solution in hexane) was added dropwise and the mixture wasstirred at −60° C. for a further 20 min Triisopropylborate (1.8 ml) wasadded before allowing the mixture to warm to room temperature. Afterstirring at room temperature for 3.5 h, the THF was removed under vacuumand DME (20 ml), water (2 ml) and2-(aminocarbonyl)amino-5-bromo-thiophene-3-carboxamide (0.47 g) wereadded. The mixture was degassed by purging with argon and sonicating for15 min, sodium hydrogen carbonate (0.445 g) and Pd(Ph₃)₄ (0.1 g) addedand heated to 90° C. for 18 h under argon. The solvent was removed undervacuum and the residue was partitioned between 2N sodium hydroxidesolution and DCM. The combined phases were filtered and the aqueousphase was taken to pH 7 by addition of 5N aqueous hydrochloric acid. Aprecipitate formed which was purified by preparative hplc to yield theproduct as a solid (0.01 g).MS (M+H)⁺407.¹H NMR (DMSO-D6) 2.62 (t, 4H), 3.19 (s, 6H), 3.39 (t, 4H), 3.61 (s, 2H),6.87-6.95 (bs, 2H), 7.22-7.31 (m, 3H), 7.44 (d, 2H), 7.61-6.9 (m, 2H),10.97 (s,1H).

EXAMPLE 42-[(Aminocarbonyl)amino]-5-{4-[(N-ethyl-N-(2-methoxyethyl)amino)methyl]phenyl}thiophene-3-carboxamide

a) The title compound was made fromN-(4-bromobenzyl)-N-(2-ethoxyethyl)ethanamine in a similar manner toExample 3 (b) except that it was purified by ion exchange chromatographyto yield a cream solid (0.18 g).

MS (M+1)⁺ 377.

¹H NMR (DMSO-D6) 0.97 (t, 3H), 2.48 (q, 2H obscured), 2.57 (t, 2H), 3.2(s, 3H), 3.39 (t, 2H), 3.55 (s, 2H), 6.91 (bs, 2H), 7.21-7.31 (m, 3H),7.44 (d, 2H), 7.62-7.69 (m, 2H), 10.97 (bs, 1H).

b) N-(4-Bromobenzyl)-N-(2-methoxyethyl)ethanamine

4-Bromobenzylbromide (4.01 g) was added to a solution ofN-(2-ethoxyethyl)ethanamine (3.64 g) in DMF (40 ml). After stirring atambient temperature for 18 h, the mixture was partitioned betweendiethyl ether and water. The organic phase was extracted with 2N aqueoushydrochloric acid which was then taken to pH 10 by addition of 2N sodiumhydroxide solution. This was then extracted with diethyl ether which wasdried and the solvent removed under vacuum to yield the title compoundas a yellow oil (4.25 g).¹H NMR (DMSO-D6) 0.95 (t, 2H), 2.45 (q, 3H), 2.55 (t, 2H), 3.2 (s, 3H),3.36 (t, 2H), 3.52 (s, 2H), 7.24 (d, 2H), 7.46 (d, 2H).

EXAMPLE 52-[(Aminocarbonyl)amino]-5-{4-[dimethylaminomethyl]phenyl}thiophene-3-carboxamide

a) The title compound was prepared from 4-(dimethylaminomethyl)phenylbromide (0.91 g) by the same method as for Example 4 (a) to give theproduct as a solid (0.155 g).

MS (M+H)⁺ 319.

¹HNMR (DMSO-D6) 2.17 (s, 6H), 3.4 (s, 2H), 7.93 (bs, 2H), 7.21-7.31 (m,3H), 7.46 (d, 2H), 7.6-7.7 (m, 2H), 10.97 (bs, 1H).

b) 4-(Dimethylaminomethyl)phenyl bromide

2M Ethanolic dimethylamine solution (30 ml) was added to a solution of4-bromobenzyl bromide (5 g) in DMF (20 ml). The mixture was stirred atambient temperature for 18 h and was then partitioned between ethylacetate and saturated aqueous sodium hydrogen carbonate solution. Theorganic phase was dried, evaporated and purified by columnchromatography, eluting with a gradient of 0 to 5% methanol in DCM toyield the product as a clear oil.MS (M+H)⁺ 214.¹H NMR (CDCl₃) 2.21 (s, 6H), 3.35 (s, 2H), 7.18 (d, 2H), 7.43 (d, 2H).

EXAMPLE 62-[(Aminocarbonyl)amino]-5-{4-[(N-(2,2,2-trifluoroethyl)-N-(2-methoxyethyl)amino)methyl]phenyl}thiophene-3-carboxamide

a) The title compound was made in a similar manner to Example 5 (a), butusing (4-bromobenzyl)-(2-methoxyethyl)-(2,2,2-trifluoroethyl)amine.

MS(ES) 429 (M−H)⁻.

¹H NMR (DMSO-D6) δ 2.7 (t, 2H), 3.2 (s, 3H), 3.2-3.4 (m, 4H), 3.75 (s,2H), 6.9 (bs, 2H), 7.2 (bs, 1H), 7.3 (d, 2H), 7.45 (d, 2H), 7.6-7.7(s+bs, 2H), 10.9 (s, 1H).

b) (4-Bromobenzyl)-(2-methoxyethyl)-(2,2,2-trifluoroethyl)amine

To a solution of 2-bromo-1,1,1-trifluoroethane (0.74 ml) andtriethylamine (1.13 ml) in DMF (30 ml) was added(4-bromobenzyl)-(2-methoxyethyl)amine (2.05 g). The resulting mixturewas stirred for 18 h. The mixture was partitioned between ethyl acetateand water, the organic layer was dried and evaporated to an oil, whichwas purified by column chromatography eluting with a gradient of ethylacetate (60 to 100%) in isohexane to give(4-bromobenzyl)-(2-methoxyethyl)-(2,2,2-trifluoroethyl) amine (1.84 g).¹H NMR (DMSO-D6) δ 2.1 (bs, 2H), 2.6 (t, 2H), 3.2 (s, 3H), 3.4 (t, 2H),3.65 (d, 2H), 7.25 (d, 2H), 7.45 (d, 2H).c) (4-Bromobenzyl)-(2-methoxyethyl)amine2-Methoxyethylamine (16 ml) was added to a solution of4-bromobenzaldehyde (17.2 g) in THF (150 ml). Glacial acetic acid (5.32ml) and MgSO₄ (0.5 g) were added and the mixture was stirred for 45 min.Sodium triacetoxyborohydride (29.7 g) was added and the resultingmixture was stirred for 4 h. The mixture was partitioned between ethylacetate and saturated sodium bicarbonate. The organic layer was driedand concentrated to an oil, which was purified by column chromatography,eluting with a gradient of ethyl acetate (10 to 100%) in isohexane, theneluting with methanol/DCM (10:90) to give(4-bromobenzyl)-(2-methoxyethyl)amine (13.0 g) as a yellow oil.MS (ES) 244 (M+H)⁺.¹H NMR (DMSO-D6) δ 2.05 (bs, 1H), 2.6 (t, 2H), 3.38 (t, 2H), 3.2 (s,3H), 3.62 (s, 2H), 7.25 (d, 2H), 7.45 (d, 2H).

EXAMPLE 72-[(Aminocarbonyl)amino]-5-[4-[(2-methoxyethyl)amino)methyl]phenyl}thiophene-3-carboxamide

a) 2-[(Aminocarbonyl)amino]-5-[4-formylphenyl]thiophene-3-carboxamide

2-[(Aminocarbonyl)amino]-5-bromothiophene-3-carboxamide (11.75 g) wasstirred in DME (500 ml) and saturated aqueous sodium bicarbonatesolution (100 ml), and 4-formylphenyl boronic acid (10 g) was added. Theflask was flushed with argon andtetrakis-(triphenylphosphine)palladium(0) (5.1 g) was then added. Thereaction was stirred at 90° C. for 2 h, then cooled and evaporated underreduced pressure. The residue was treated with DCM (200 ml) and 2Nsodium hydroxide solution (100 ml), and stirred for 20 min. Theresulting solid was then isolated by filtration and purified bytrituration with ethanol (100 ml), giving the product as a pale greensolid (5.75 g).

MS (ES) 290 (M+H)⁺.

¹H NMR (DMSO-D6) 7.05 (s, 2H), 7.40 (s, 1H), 7.75 (m, 3H), 7.90 (d, 2H),8.00 (s, 1H), 9.95 (s, 1H), 11.10 (s, 1H).

b)2-[(Aminocarbonyl)amino]-5-{4-[(2-methoxyethyl)amino)methyl]phenyl}thiophene-3-carboxamide2-[(Aminocarbonyl)amino]-5-[4-formylphenyl]thiophene-3-carboxamide (300mg) was stirred for 18 h in a mixture of methanol (10 ml),(2-methoxyethyl)amine (0.18 ml) and sodium cyanoborohydride (200 mg). 4NHydrochloric acid was added until the mixture was acid. It was filtered,evaporated to dryness, dissolved in 2N sodium hydroxide (5 ml), filteredand the pH adjusted to 7 with solid potassium carbonate. The resultingsolid was filtered off, washed with water, ether and dried to give theproduct as a fawn solid (180 mg).

MS (ES) 349 (M+H)⁺.

¹H NMR (DMSO-D6) 2.76 (t, 2H), 3.24 (s, 3H), 3.44 (t, 2H), 3.81 (s, 2H),6.93 (bs, 2H), 7.30 (bs, 1H), 7.36 (d, 2H), 7.48 (d, 2H), 7.66 (bs, 1H),7.71 (s, 1H), 10.98 (s, 1H).

EXAMPLE 82-[(Aminocarbonyl)amino]-5-[4-{[(2-(1H-indol-3-yl)ethyl)amino]methyl}phenyl)-thiophene-3-carboxamide

2-[(Aminocarbonyl)amino]-5-(4-formylphenyl)thiophene-3-carboxamide (0.05g) was stirred in a mixture of DME (15 ml), methanol (15 ml) and glacialacetic acid (0.1 ml). Tryptamine (0.136 g) was added and the reactionwas stirred at 80° C. for 2 h, and then polymer-supportedcyanoborohydride (0.115 g) was added. The reaction was stirred at 80° C.for a further 4 h, and then polymer-supported benzaldehyde (1.33 g) wasadded. The resins were removed by filtration, and the filtrate was thenpassed through a 10 g SCX column, washing with methanol (100 ml). Theproduct was eluted using 1M methanolic ammonia (50 ml), this solutionwas then evaporated to dryness under reduced pressure and the residuepurified by chromatography on silica, eluting with DCM/methanol (2M NH₃)(20:1), to give the product as an yellow powder (0.032 g).MS (ES) 434 (M+H)⁺.¹H NMR (DMSO-D6) 3.00 (s, 4H), 3.96 (s, 2H), 6.95-7.18 (m, 4H),7.10-7.15 (m, 1H), 7.21 (d, 1H), 7.35 (s, 1H), 7.40 (d, 1H), 7.47 (d,2H), 7.54-7.59 (m, 3H), 7.73 (s, 1H), 7.79 (s, 1H), 11.05 (s, 1H).The compounds of Examples 9 to 36 were prepared from2-[(aminocarbonyl)amino]-5-(4-formylphenyl)thiophene-3-carboxamide andthe appropriate amine using the general method of Example 8.

EXAMPLE 92-[(Aminocarbonyl)amino]-5-[4-{methylaminomethyl}phenyl]thiophene-3-carboxamide

MS (ES) 303 (M−H)⁻.

¹H NMR (DMSO-D6) 2.29 (s, 3H), 3.65 (s, 2H), 6.94 (s, 2H), 7.27 (s, 1H),7.34 (d, 2H), 7.48 (d, 2H), 7.66 (s, 1H), 7.70 (s, 1H), 11.00 (s, 1H).

EXAMPLE 102-[(Aminocarbonyl)amino]-5-[4-{cyclopropylaminomethyl}phenyl]thiophene-3-carboxamide

MS (ES) 329 (M−H)⁻.

¹H NMR (DMSO-D6) 0.23-0.28 (m, 2H), 0.32-0.38 (m, 2H), 2.06 (m, 1H),3.71 (s, 2H), 6.94 (s, 2H),7.27 (s, 1H), 7.34 (d, 2H), 7.46 (d, 2H),7.63-7.73 (m, 2H), 11.00 (s, 1H).

EXAMPLE 112-[(Aminocarbonyl)amino]-5-[4-{([2R)-2-hydroxypropyl]amino)methyl}phenyl]thiophene-3-carboxamide

MS (ES) 349 (M+H)⁺.

¹H NMR (DMSO-D6) 1.05 (d, 3H), 2.42 (d, 2H), 3.70 (s, 2H), 4.42 (m, 1H),6.94 (s, 2H), 7.27 (s, 1H), 7.34 (d, 2H), 7.47 (d, 2H), 7.64-7.71 (m,2H), 11.00 (s, 1H).

EXAMPLE 122-[(Aminocarbonyl)amino]-5-[4-{([(2S)-2-hydroxypropyl]amino)methyl}phenyl]thiophene-3-carboxamide

MS (ES) 349 (M+H)⁺.

¹H NMR (DMSO-D6) 1.05 (d, 3H), 2.42 (d, 2H), 3.70 (s, 2H), 4.42 (m, 1H),6.94 (s, 2H), 7.27 (s, 1H), 7.34 (d, 2H), 7.47 (d, 2H), 7.64-7.71 (m,2H), 11.00 (s, 1H).

EXAMPLE 132-[(Aminocarbonyl)amino]-5-[4-{[(tetrahydrofuran-2-ylmethyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 375 (M+H)⁺.

¹H NMR (DMSO-D6) 1.47-1.58 (m, 1H), 1.74-1.94 (m, 3H), 2.52-2.56 (m,2H), 3.56-3.63 (m, 1H), 3.68-3.77 (m, 3H), 3.84-3.92 (m, 1H), 6.94 (s,2H), 7.27 (s, 1H), 7.33 (d, 2H), 7.47 (d, 2H), 7.66 (s, 1H), 7.69 (s,1H), 11.00 (s, 1H).

EXAMPLE 142-[(Aminocarbonyl)amino]-5-[4-{[(4-fluorobenzyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 399 (M+H)⁺.

¹H NMR (DMSO-D6) 3.69 (bs, 4H), 6.94 (s, 2H), 7.10-7.14 (m, 2H), 7.28(s, 1H), 7.34-7.42 (m, 4H), 7.47 (d, 2H), 7.66 (s, 1H), 7.70 (s, 1H),11.00 (s, 1H).

EXAMPLE 152-[(Aminocarbonyl)amino]-5-[4-{[(3-{2-oxopyrrolidin-1-yl}propyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 416 (M+H)⁺.

¹H NMR (DMSO-D6) 1.56-1.67 (m, 2H), 1.85-1.95 (m, 2H), 2.19 (t, 2H),2.45 (2H obscured), 3.21 (t, 2H), 3.30 (2H obscured), 3.68 (s, 2H), 6.94(s, 2H), 7.28 (s, 1H), 7.33 (d, 2H), 7.47 (d, 2H), 7.67 (s, 1H), 7.70(s, 1H), 10.99 (s, 1H).

EXAMPLE 162-[(Aminocarbonyl)amino]-5-[4-{[(1-naphthylmethyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 431 (M+H)⁺.

¹H NMR (DMSO-D6) 3.81 (s, 2H), 4.14 (s, 2H), 6.94 (s, 2H), 7.28 (s, 1H),7.39-7.57 (m, 8H), 7.69 (s, 1H), 7.71 (s, 1H), 7.83 (d, 1H), 7.90-7.94(m, 1H), 8.13-8.18 (m, 1H), 10.99 (s, 1H)

EXAMPLE 172-[(Aminocarbonyl)amino]-5-[4-{([1-(4-chlorobenzyl)-2-hydroxyethyl]amino)methyl}phenyl]thiophene-3-carboxamide

MS (ES) 459 (M+H)⁺.

¹H NMR (DMSO-D6) 2.66-2.75 (m, 3H), 3.30 (1H obscured), 3.74 (s, 2H),4.54 (m, 1H), 6.94 (s, 2H), 7.21-7.35 (in 7H), 7.44 (d, 2H), 7.63-7.70(m, 2H), 10.99 (s, 1H).

EXAMPLE 182-[(Aminocarbonyl)amino]-5-[4-{(cyclopentylamino)methyl}phenyl]thiophene-3-carboxamide

MS (ES) 359 (M+H)⁺.

¹H NMR (DMSO-D6) 1.28-1.79 (m, 8H), 3.01 (s, 1H), 3.67 (s, 2H), 6.94 (s,2H), 7.27 (s, 1H), 7.34 (d, 2H), 7.46 (d, 2H), 7.62-7.73 (m, 2H), 10.99(s, 1H).

EXAMPLE 192-[(Aminocarbonyl)amino]-5-[4-{[(pyridin-3-ylmethyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 382 (M+H)⁺.

¹H NMR (DMSO-D6) 3.69 (s, 2H), 3.72 (s, 2H), 6.94 (s, 2H), 7.27 (s, 1H),7.32-7.40 (m, 3H), 7.48 (d, 2H), 7.67 (s, 1H), 7.70 (s, 1H), 7.77 (d,1H), 8.45 (d, 1H), 8.54 (s, 1H) 11.00 (s, 1H).

EXAMPLE 202-[(Aminocarbonyl)amino]-5-[4{[(2-[pyridin-2-yl]ethyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 396 (M+H)⁺.

¹H NMR (DMSO-D6) 2.88-2.92 (m, 4H), 3.76 (s, 2H), 6.94 (s, 2H),7.17-7.22 (A, 1H), 7.27 (d, 2H), 7.33 (d, 2H), 7.46 (d, 2H), 7.64-7.71(m, 3H), 8.47 (m, 1H). 11.00 (s, 1H).

EXAMPLE 212-[(Aminocarbonyl)amino]-5-[4-{[({2-hydroxy-1,1-dimethyl}ethyl)amino]methyl)}phenyl]thiophene-3-carboxamide

MS (ES) 363 (M+H)⁺.

¹H NMR (DMSO-D6) 1.02 (s, 6H), 3.30 (2H obscured), 3.64 (s, 2H), 6.92(s, 2H), 7.26 (s, 1H), 7.35 (d, 2H), 7.46 (d, 2H), 7.68 (m, 2H), 11.00(s, 1H).

EXAMPLE 222-[(Aminocarbonyl)amino]-5-[4-{[({1,2-diphenyl}ethyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 471M+H)⁺.

¹H NMR (DMSO-D6) 2.78-2.88 (m, 1H), 2.94-3.03 (m, 1H), 3.40 (d, 1H),3.54 (d, 1H), 3.76-3.85 (m, 1H), 6.94 (s, 2H), 7.04-7.09 (m, 2H),7.12-7.32 (m, 11H), 7.40 (d, 2H), 7.62-7.70 (m, 2H), 11.00 (s, 1H).

EXAMPLE 232-[(Aminocarbonyl)amino]-5-[4-{[({2-methoxy-1-methyl}etyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 363 (M+H)⁺.

¹H NMR (DMSO-D6) 0.99 (d, 3H), 2.80 (m, 1H), 3.16-3.27 (m, 5H),3.65-3.81 (m, 2H), 6.94 (s, 2H), 7.28 (s, 1H), 7.34 (d, 2H), 7.47 (d,2H) 7.63-7.72 (m, 2H), 11.00 (s, 1H).

EXAMPLE 242-[(Aminocarbonyl)amino]-5-[4-{[({2-hydroxy-1-methyl}ethyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 349 (M+H)⁺.

¹H NMR (DMSO-D6) 0.96 (d, 3H), 3.30 (2H obscured), 3.64-3.80 (m, 2H),4.48 (m, 1H), 6.94 (s, 2H), 7.27 (s, 1H), 7.35 (d, 2H), 7.47 (d, 2H),7.64-7.72 (m, 2H), 11.00 (s, 1H).

EXAMPLE 252-[(Aminocarbonyl)amino]-5-[4-1[(2-methylbenzyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 395 (M+H)⁺.

¹H NMR (MSO-D6) 2.27 (s, 3H), 3.66 (s, 2H), 3.74 (s, 2H), 6.95 (s, 2H),7.12-7.17 (m, 3H), 7.28 (s, 1H), 7.31-7.36 (m, 1H), 7.39 (d, 2H), 7.49(d, 2H), 7.67 (s, 1H), 7.70 (s, 1H), 11.00 (s, 1H).

EXAMPLE 262-[(Aminocarbonyl)amino]-5-[4-{[(3-methoxybenzyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 411 (M+H)⁺.

¹H NMR (DMSO-D6) 3.67 (s, 4H), 3.75 (s, 3H), 6.77-6.82 (m, 1H),6.88-6.98 (m, 4H), 7.22 (t, 1H), 7.27 (s, 1H), 7.36 (d, 2H), 7.47 (d,2H), 7.67 (s, 1H), 7.70 (s, 1H), 11.00 (s, 1H).

EXAMPLE 272-[(Aminocarbonyl)amino]-5-[4-{[(2-fluorobenzyl)amino]methyl}pheny]thiophene-3-carboxamide

MS (ES) 399M+H)⁺.

¹H NMR (DMSO-D6) 3.71 (s, 2H), 3.73 (s, 2H), 6.94 (s, 2H), 7.10-7.22 (m,2H), 7.24-7.33 (m, 2H), 7.37 (d, 2H), 7.45-7.55 (m, 3H), 7.67 (s, 1H),7.70 (s, 1H), 11.00 (s, 1H).

EXAMPLE 282-[(Aminocarbonyl)amino]-5-[4-{[3-fluorobenyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 399 (M+H)⁺.

¹H NMR (DMSO-D6) 3.68 (s, 2H), 3.72 (s, 2H), 6.94 (s, 2H), 7.01-7.08 (m,1H), 7.15-7.23 (m, 2H), 7.28 (s, 1H), 7.32-7.40 (m, 3H), 7.48 (d, 2H),7.67 (s, 1H), 7.70 (s, 1H), 11.00 (s, 1H).

EXAMPLE 292-[(Aminocarbonyl)amino]-5-[4-{[(4-{Phenyl}butyl)amino]methyl}pheynl]thiophene-3-carboxamide

MS (ES) 421 (M−H)⁻.

¹H NMR (DMSO-D6) 1.38-1.50 (m, 2H), 1.55-1.66 (m, 2H), 2.51 (2Hobscured), 2.57 (t, 2H), 3.66 (s, 2H), 6.94 (s, 2H), 7.12-7.21 (m, 3H),7.23-7.29 (m, 3H), 7.33 (d, 2H), 7.46 (d, 2H), 7.62-7.74 (m, 2H), 11.00(s, 1H).

EXAMPLE 302-[(Aminocarbonyl)amino]-5-[4-([3-(trifluoromethyl)benzyl]amino)methyl}phenyl]thiophene-3-carboxamide

MS (ES) 447 (M−H)⁻.

¹H NMR (DMSO-D6) 3.69 (s, 2H), 3.79 (s, 2H), 6.95 (s, 2H), 7.28 (s, 1H),7.36 (d, 2H), 7.48 (d, 2H), 7.53-7.75 (m, 6H), 11.00 (s, 1H).

EXAMPLE 312-[(Aminocarbonyl)amino]-5-[4-{[(5-cyanonentyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 384 (M−H)⁻.

¹H NMR (DMSO-D6) 1.34-1.50 (m, 6H), 1.51-1.60 (m, 2H), 2.48 (t, 2H),3.67 (s, 2H), 6.94 (s, 2H), 7.28 (s, 1H), 7.34 (d, 2H), 7.46 (d, 2H),7.61-7.73 (m, 2H), 11.00 (s, 1H).

EXAMPLE 322-[(Aminocarbonyl)amino]-5-[4-{({2-methyl}propylamino)methyl}phenyl]thiophene-3-carboxamide

The product was purified by reverse phase preparative HPLC yielding thetrifluoroacetate salt as a white powder (0.055 g).

MS (ES) 347 (M+H)⁺.

¹H NMR (DMSO-D6) 0.95 (d, 6H), 1.93-2.02 (m, 1H), 2.75-2.82 (m, 2H),4.14 (s, 2H), 6.99 (s, 2H), 7.33 (s, 1H), 7.52 (d, 2H), 7.60 (d, 2H),7.68 (s, 1H), 7.81 (s, 1H), 8.7 (s, 2H), 11.0 (s, 1H).

EXAMPLE 332-[(Aminocarbonyl)amino]-5-[4-{[(4-methoxybenzyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 411 (M+H)⁺.

¹H NMR (DMSO-D6) 3.65 (bs, 4H), 3.75 (s, 3H), 6.88 (d, 2H), 6.94 (s,2H), 7.22-7.31 (m, 3H), 7.35 (d, 2H), 7.47 (d, 2H), 7.66 (s, 1H), 7.69(s,1H), 11.00 (s, 1H).

EXAMPLE 342-[(Aminocarbonyl)amino]-5-[4-{[(2-phenylethyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 395 M+H)⁺.

¹H NMR (DMSO-D6) 2.74 (s, 4H), 3.72 (s, 2H), 6.94 (s, 2H), 7.14-7.35 (m,8H), 7.46 (d, 2H), 7.67 (s, 1H), 7.70 (s, 1H), 11.00 (s, 1H).

EXAMPLE 352-[(Aminocarbonyl)amino]-5-[4-{[(2-hydroxyethyl)amino]methyl}phenyl]thiophene-3-carboxamide

The product was purified by reverse phase preparative HPLC yielding thetrifluoroacetate salt as a white powder.

MS (ES) 335 (M+H)⁺.

¹H NMR (DMSO-D6) 2.99 (t, 2H), 3.64-3.70 (m, 2H), 4.17 (s, 2H), 5.22 (t,1H), 6.99 (s, 2H), 7.33 (s, 1H), 7.52 (d, 2H), 7.59 (d, 2H), 7.68 (s,1H), 7.80 (s, 1H), 8.83 (s, 2H), 11.02 (s, 1H).

EXAMPLE 362-[(Aminocarbonyl)amino]-5-[4-{[({2-methoxy-2-methyl}propyl)amino]methyl}phenyl]thiophene-3-carboxamide

MS (ES) 377 (M+H)⁺.

¹H NMR (DMSO-D6) 1.11 (s, 6H), 2.37-2.48 (m, 2H), 3.07 (s, 3H), 3.72 (s,2H), 6.97 (s, 2H), 7.26-7.40 (m, 3H), 7.44-7.52 (m, 2H), 7.65-7.76 (m,2H), 11.00 (s, 1H).

EXAMPLE 373-[(Aminocarbonyl)amino]-5-[4-{[N-(4-fluorobenzyl)-N-methylamino]methyl}phenyl]thiophene-2-carboxamide

a) 2-Bromothiophene-4-carboxylic acid

Prepared according to the method as described in J. Am. Chem. Soc.,1954, 76,2445.

MS (ES) 205 (M−H)⁻.

¹H NMR (DMSO-D6) 7.45 (s, 1H), 8.22 (s, 1H), 12.94 (bs, 1H).

b) 2-Bromo-4 (N-t-butyloxycarbonyl)aminothiophene

2-Bromothiophene-4-carboxylic acid (3 g) was dissolved in dry warmt-butanol (24 ml). Triethylamine (2.02 ml) was added followed bydiphenylphosphoryl azide (3.12 ml). The solution was heated slowly toreflux and heating continued at reflux overnight. The reaction mixturewas then allowed to cool, poured into water (150 ml) and extracted withethyl acetate (3×100 ml). The combined extracts were dried, filtered andevaporated. The crude product was purified by column chromatography,eluting with 5% ethyl acetate in hexane, to give a white solid (1.69 g).MS (ES) 276 (M−H)⁻.¹H NMR (DMSO-D6) 1.44 (s, 9H), 7.03 (s, 1H), 7.51 (s, 1H), 9.65 (s, 1H).c) 5-Bromo-3-[(t-but loxycarbonyl)amino]thiophene-2-carboxylic acid2-Bromo-4-(N-t-butyloxycarbonyl)aminothiophene (1.68 g) was stirred indry THF (45 ml) under argon and the solution was cooled to −78° C.Lithium diisopropylamide (7.55 ml, 2M solution) was added dropwise andstirring continued for 3.5 h. Powdered CO₂ (excess) was added and themixture stirred for a further 10 minutes before allowing to warm to roomtemperature. Water (50 ml) was added, the THF was removed in vacuo andthe aqueous phase was extracted with ethyl acetate (3×40 ml). Thecombined extracts were washed with 1M hydrochloric acid (50 ml), water(50 ml) and brine (50 ml), dried, filtered and the solvent evaporated.The residue was triturated with DCM and the product collected byfiltration as a pale yellow solid (1.57 g).MS (ES) 320 (M−H)⁻.¹H NMR (DMSO-D6) 9.38 (s, 1H), 7.79 (s, 1H), 1.42 (s, 9H).d) 5-Bromo-3-(t-butyloxycarbonyl)aminothiophene-2-carboxamide5-Bromo-3-[(t-butyloxycarbonyl)amino]thiophene-2-carboxylic acid (0.80g) was stirred in acetonitrile (80 ml). Hydroxybenztriazole (1.41 g) and1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (2.62 g)were added and stirring continued at room temperature for 10 min.Concentrated aqueous ammonia solution (8 ml) was added and the reactionmixture was heated to reflux for 1 h. The acetonitrile was removed byevaporation. Water (100 ml) was added and the mixture was sonicated andtriturated. The resultant off-white solid was then collected byfiltration, washed with water and dried under vacuum (0.763 g).MS (ES) 319 (M−H)³¹.¹H NMR (DMSO-D6) 1.45 (s, 9H), 7.63 (brs, 2H), 7.78 (s, 1H), 10.40 (s,1H).e) 3-Amino-5-bromothiophene-2-carboxamide5-Bromo-3-(t-butyloxycarbonyl)aminothiophene-2-carboxamide (0.76 g) wasstirred in DCM (30 ml). Trifluoroacetic acid (5 ml) was added, thesolution was stirred at room temperature for 1 h, poured into saturatedaqueous sodium hydrogen carbonate solution (200 ml) and extracted withDCM (3×100 ml). The combined extracts were washed with brine (150 ml),dried, filtered and evaporated to give a yellow solid (0.511 g).MS (ES) 221 (M+H)⁺.¹H NMR (DMSO-D6) 6.50 (bs, 2H), 6.69 (s, 1H), 6.87 (bs, 2H).f) 3-[(Aminocarbonyl)amino-5-bromothiophene-2-carboxamide3-Amino-5-bromothiophene-2-carboxamide (0.25 g) was stirred in anhydrousTHF (10 ml), cooled to 0° C. and trichloroacetylisocyanate (0.148 ml)added dropwise. The mixture was allowed to warm to ambient temperature,stirred for 1.5 h and 2M ammonia in methanol (16 ml) added. After 1.5 h,the solvents were evaporated and the residue triturated with diethylether and dried in vacuo to give the title compound as a yellow solid(0.26 g).MS (ES) 264 (M+H)⁺.¹H NMR (DMSO-D6) 6.63 (bs, 2H), 7.41 (bs, 2H), 7.97 (s, 1H), 10.02 (s,1H).g) 3-[(Aminocarbonyl)amino]-5-[4-formylphenyl]thiophene-2-carboxamide3-[(Aminocarbonyl)amino]-5-bromothiophene-2 carboxamide (2 g) wasstirred in DME (200 ml) and saturated aqueous sodium bicarbonatesolution (40 ml), and 3-formylphenyl boronic acid (1.7 g) was added. Theflask was flushed with argon, andtetrakis-(triphenylphosphine)palladium(0) (0.878 g) was then added Thereaction was stirred at 90° C. for 2 h, then cooled and evaporated underreduced pressure. The residue was treated with DCM (200 ml) andsaturated aqueous sodium bicarbonate solution (100 ml) and stirred for20 min. The resulting solid was then isolated by filtration, andpurified by sonication in ethanol (100 ml) and the solid isolated byfiltration, giving the product as an off white solid (1.53 g).MS (ES) 288 (M−H)⁻.¹H NMR (DMSO-D6) 6.68 (s, 2H), 7.52 (s, 2H), 7.67-7.75 (m, 1H),7.90-7.98 (m, 2H), 8.13 (s, 1H), 8.37 (s, 1H), 10.03-10.12 (m, 2H).h)3-[(Aminocarbonyl)amino]-5-[4-{[N-(4-fluorobenzyl)-N-methylamino]methyl}phenyl]thiophene-2-carboxamideThe title compound was made from3-[(aminocarbonyl)amino]-5-[4-formylphenyl]thiophene-2-carboxamide usingthe general method of Example 8.MS (ES) 413 (M+H)⁺.¹H NMR (DMSO-D6) 2.09 (s, 3H), 3.48-3.56 (m, 4H), 6.64 (s, 2H),7.12-7.21 (m, 2H), 7.34-7.50 (m, 6H), 7.59 (s, 2H), 8.24 (s, 1H), 10.07(s, 1H).The compounds of Examples 38 to 55 were prepared from3-[(aminocarbonyl)amino]-5-(4-formylphenyl) thiophene-2-carboxamide andthe appropriate amine using the general method of Example 8.

EXAMPLE 383-[(Aminocarbonyl)amino]-5-[4-{[N-(2-{pyridin-2-yl}ethyl)methylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 410 (M+H)⁺.

¹H NMR (DMSO-D6) 2.21 (s, 3H), 2.72 (t, 2H), 2.93 (t, 2H), 3.56 (s, 2H),6.63 (s, 2H), 7.17-7.22 (m, 1H), 7.26-7.34 (m, 3H), 7.43 (s, 2H), 7.54(d, 2H), 7.66-7.73 (m, 1H), 8.22 (s, 1H), 8.46 (d, 1H), 10.06 (s, 1H).

EXAMPLE 393-[(Aminocarbonyl)amino]-5-[4-{[N-(pyridin-2-ylmethyl)methylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 396 (M+H)⁺.

¹H NMR (DMSO-D6) 2.10 (s, 3H), 3.56 (s, 4H), 6.64 (s, 2H), 7.36-7.50 (m,5H), 7.60 (d, 2H), 7.77 (d, 1H), 8.24 (s, 1H), 8.45-8.59 (m, 2H), 10.06(s, 1H).

EXAMPLE 403-[(Aminocarbonyl)amino]-5-[4-{[N-benzyl-N-methylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 395 (M+H)⁺.

¹H NMR (DMSO-D6) 2.10 (s, 3H), 3.49-3.57 (m, 4H), 6.60 (s, 2H),7.22-7.29 (m, 1H), 7.31-7.48 (m, 8H), 7.60 (d, 2H), 8.23 (s, 1H), 10.06(s, 1H).

EXAMPLE 413-[(Aminocarbonyl)amino]-5-[4-{[N-(2-methoxyethyl)ethylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 377 (M+H)⁺.

¹H NMR (DMSO-D6) 0.99 (t, 3H), 2.50 (2H, obscured), 2.61 (t, 2H), 3.23(s, 3H), 3.43 (t, 2H), 3.62 (s, 2H), 6.60 (s, 2H), 7.34-7.47 (m, 4H),7.57 (d, 2H), 8.23 (s, 1H), 10.06 (s, 1H).

EXAMPLE 423-[(Aminocarbonyl)amino]-5-[4-{[(N-2-methoxyethyl})methylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 363 (M+H)⁺.

¹H NMR (DMSO-D6) 2.18 (s, 3H), 2.53-2.59 (m, 2H), 3.24 (s, 3H), 3.47 (t,2H), 3.53 (s, 2H), 6.60 (s, 2H), 7.34-7.48 (m, 0.4H), 7.58 (d, 2H), 8.23(s, 1H), 10.06 (s, 1H).

EXAMPLE 433-[(Aminocarbonyl)amino]-5-[4-{[N-benzyl-N-(2-cyanoethyl)amino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 434 (M+H)⁺.

¹H NMR (DMSO-D6) 2.63-2.77 (m, 4H), 3.60-3.67 (m, 4H), 6.60 (s, 2H),7.23-7.29 (m, 1H), 7.32-7.46 (m, 6H), 7.48 (d, 2H), 7.60 (d, 2H), 8.23(s, 1H), 10.06 (s, 1H).

EXAMPLE 443-[(Aminocarbonyl)amino]-5-[4-{[N-(2-cyanoethyl)-N-methylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 358 (M+H)⁺.

¹H NMR (DMSO-D6) 2.19 (s, 3H), 2.62-2.74 (m, 4H), 3.57 (s, 2H), 6.59 (s,2H), 7.35-7.46 (m, 4H), 7.59 (d, 2H), 8.23 (s, 1H), 10.06 (s, 1H).

EXAMPLE 453-[(Aminocarbonyl)amino]-5-[4-{[N-isopropyl-N-(2-methoxyethyl)amino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 391 (M+H)⁺.

¹H NMR (DMSO-D6) 0.99 (d, 6H), 2.57 (t, 2H), 2.85-2.93 (m, 1H), 3.18 (s,3H), 3.30 (2H obscured), 3.60 (s, 2H), 6.60 (s, 2H), 7.34-7.45 (m, 4H),7.56 (d, 2H), 8.21 (s, 1H), 10.06 (s, 1H).

EXAMPLE 463-[(Aminocarbonyl)amino]-5-[4-{[(3-methoxybenzyl)amino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 411 (M+H)⁺.

¹H NMR (DMSO-D6) 3.65-3.79 (m, 7H), 6.60 (s, 2H), 6.77-6.83 (m, 1H),6.89-6.98 (m, 2H), 7.23 (t, 1H), 7.39 (s, 2H), 7.44 (d, 2H), 7.58 (d,2H), 8.23 (s, 1H), 10.06 (s, 1H).

EXAMPLE 473-[(Aminocarbonyl)amino]-5-[4-{[N-(2-methoxyethyl)-N-(pyridin-2-yl)methylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 440 (M+1)⁺.

¹H NMR (DMSO-D6) 2.66 (t, 2H), 3.20 (s, 3H), 3.46 (t, 2H), 3.70 (s, 2H),3.76 (s, 2H), 6.60 (s, 2H), 7.22-7.28 (m, 1H), 7.39 (s, 2H), 7.43-7.61(m, 5H), 7.74-7.82 (m, 1H), 8.23 (s, 1H), 8.46-8.51 (m, 1H), 10.06 (s,1H).

EXAMPLE 483-[(Aminocarbonyl)amino]-5-[4-{[N-(3,5-dimethyl-1H-pyrazol-4-yl)methyl]methylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 413 (M+H)⁺.

¹H NMR (DMSO-D6) 2.01 (s, 3H), 2.07-2.16 (m, 6H), 3.23 (s, 2H), 3.44 (s,2H), 6.60 (s, 2H), 7.33-7.44 (m, 4H), 7.58 (d, 2H), 8.23 (s, 1H), 10.06(s, 1H).

EXAMPLE 493-[(Aminocarbonyl)amino]-5-[4-{({N-[3-methylisoxazol-5-yl]methyl]methylamino)methyl}phenyl]thiophene-2-carboxamide

MS (ES) 400 (M+H)⁺.

¹H NMR (DMSO-D6) 2.18 (s, 3H), 2.23 (s, 3H), 3.57 (s, 2H), 3.71 (s, 2H),6.29 (s, 1H), 6.60 (s, 2H), 7.36-7.47 (m, 4H), 7.60 (d, 2H), 8.24 (s,1H), 10.06 (s, 1H).

EXAMPLE 503-[(Aminocarbonyl)amino]-5-[4-{[N-(2-hydroxyethyl)methylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 349. (M+H)⁺.

¹H NMR (DMSO-D6) 2.17 (s, 3H), 2.45 (t, 2H), 3.47-3.57 (m, 4H),4.34-4.39 (m, 1H), 6.60 (s, 2H), 7.32-7.46 (m, 4H), 7.57 (d, 2H), 8.23(s, 1H), 10.06 (s, 1H).

EXAMPLE 513-[(Aminocarbonyl)amino]-5-[4-{[(2-methoxyethyl)amino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 349 (M+H)⁺.

¹H NMR (DMSO-D6) 2.69 (t, 2H), 3.25 (s, 3H), 3.42 (t, 2H), 3.77 (s, 2H),6.60 (s, 2H), 7.34-7.47 (m, 4H), 7.58 (d, 2H), 8.23 (s, 1H), 10.06 (s,1H).

EXAMPLE 523-[(Aminocarbonyl)amino]-5-[4-{[N-(1-dioxidotetrahydro-3-thienyl)methylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 423 (M+H)⁺.

¹H NMR (DMSO-D6) 2.02-2.19 (m, 4H), 2.28-2.39 (m, 1H), 3.01-3.14 (m,2H), 3.21-3.38 (2H obscured), 3.44-3.69 (m, 3H), 6.60 (s, 2H), 7.33-7.47(m, 4H), 7.59 (d, 2H), 8.23 (s, 1H), 10.06 (s, 1H).

EXAMPLE 533-[(Aminocarbonyl)amino]-5-[4-{[N-benzyl-N-(2-hydroxyethyl)amino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 425 (M+H)⁺.

¹H NMR (DMSO-D6) 2.57 (t, 2H), 3.43-3.50 (m, 2H), 3.59-3.64 (m, 2H),3.70 (s, 2H), 4.35-4.40 (m, 1H), 6.60 (s, 2H), 7.27-7.42 (m, 7H), 7.46(d, 2H), 7.58 (d, 2H), 8.22 (s, 1H), 10.06 (s, 1H).

EXAMPLE 543-[(Aminocarbonyl)amino]-5-[4-{[(tetrahydrofuran-2-ylmethyl)amino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 375 (M+H)⁺.

¹H NMR (DMSO-D6) 1.47-1.59 (m, 1H), 1.73-1.95 (m, 4H), 3.55-3.77 (m,5H), 3.84-3.94 (m, 1H), 6.60 (s, 2H), 7.34-7.51 (m, 4H), 7.56 (d, 2H),8.22 (s, 1H), 10.06 (s, 1H).

EXAMPLE 55K3-[(Aminocarbonyl)amino]-5-[4-{[(2-methoxy-2-methyl)propylamino]methyl}phenyl]thiophene-2-carboxamide

MS (ES) 377 (M+H)⁺.

¹H NMR (DMSO-D6) 1.09 (s, 6H), 2.42 (s, 2H), 3.04 (s, 3H), 3.73 (s, 2H),6.60 (s, 2H), 7.31-7.44 (m, 4H), 7.54 (d, 2H), 8.19 (s, 1H), 10.03 (s,1H).

EXAMPLE 562-[(Aminocarbonyl)amino]-5-[4-{[N-(2-methoxyethyl)methylamino]methyl{phenyl]thiophene-3-carboxamide

a) N-(4-Bromobenzyl)-2-methoxy-N-methylethanamine

4-Bromobenzylbromide (1.02 g) and N-(2-methoxyethyl)methylamine (0.73 g)were stirred in DMF (20 ml) for 16 h at ambient temperature. Thereaction mixture was then partitioned between diethyl ether (40 ml) andwater (40 ml). The aqueous phase was extracted further with diethylether (40 ml) and the combined organics were washed with water (50 ml),dried, concentrated in vacuo and purified by column chromatography,eluting with a gradient of ethyl acetate (0 to 50%) in iso-hexane togive the product as a yellow oil (0.67 g).MS (ES) 258 (M+H)⁺.¹H NMR (DMSO-D6) 2.10 (s, 3H), 2.45 (m, 2H), 3.20 (s, 3H), 3.40 (m, 4H),7.21 (m, 2H), 7.45 (m, 2H).b) The title compound was prepared fromN-(4-bromobenzyl)-2-methoxy-N-methylethanamine (0.632 g) and2-(aminocarbonyl)amino-5-bromo-thiophene-3-carboxamide (0.215 g) in asimilar manner to Example 3 (b). The product was obtained directly fromthe work-up as a brown solid (0.142 g).MS (ES) 363 (M+H)⁺.¹H NMR (DMSO-D6) 2.15 (s, 3H), 2.50 (t, 2H), 3.20 (s, 3H), 3.45 (m, 4H),6.90 (s, 2H), 7.25 (m, 3H), 7.42 (d, 2H), 7.62 (m, 2H), 10.96 (s, 1H).

EXAMPLE 572-[(Aminocarbonyl)amino]-5-[4-(N,N-diethylamino)methyl]phenyl}thiohene-3-carboxamide

2-[(Aminocarbonyl)amino]-5-[4-formylphenyl]thiophene-3-carboxamide (0.1g) was stirred in a mixture of DME (10 ml) and DMA (5 ml). Diethylamine(0.2 ml) was added, followed by trimethyl orthoformate (5 ml) and aceticacid (0.5 ml). The reaction was stirred at 80° C. for 20 min, and thenpolymer-supported cyanoborohydride (0.45 g) was added. The reaction wasstirred at 80° C. for a further 2 h, and then polymer-supportedisocyanate (0.5 g) was added. The resins were removed by filtration, andthe filtrate passed through a 5 g SCX column, washing with methanol (25ml). The product was eluted using 1M methanolic ammonia (45 ml), thissolution was evaporated to dryness under reduced pressure and theresidue purified by chromatography on silica, eluting with DCM/methanol(9:1), to give the product as an off-white solid (0.012 g).MS (ES) 347 (M+H)⁺.¹H NMR DMSO-D6) 0.95 (m, 6H), 2.55 (m, 4H), 3.50 (s, 2H), 6.90 (s, 2H),7.25 (s, 1H), 7.30 (d, 2H), 7.45 (d, 2H), 7.70 (m, 2H), 11.00 (s, 1H).

EXAMPLE 582-[(Aminocarbonyl)amino]-5-[4-{([N-(2-cyanoethyl)-N-methylamino]methyl}phenyl]thiophene-3-carboxamide

The title compound was prepared using the general method of Example 57.

MS (ES) 358 (M+H)⁺.

¹H NMR (IMSO-D6) 2.20 (s, 3H), 2.60 (m, 2H), 2.75 (m, 2H), 3.55 (s, 2H),6.90 (s, 2H), 7.30 (s, 11), 7.40 (d, 2H), 7.55 (d, 2H), 7.70 (s, 1H),7.75 (s, 1H), 11.00 (s, 1H).

EXAMPLE 592-[(Aminocarbonyl)amino]-5-[4-{[(N-benzyl-N-cyanoethylamino]methyl}phenyl]thiophene-3-carboxamide

The title compound was prepared using the general method of Example 57.

MS (ES) 434 (M+H)⁺.

¹H NMR (DMSO-D6) 2.65 (m, 2H), 2.75 (m, 2H), 3.6 (s, 4H), 6.90 (s, 2H),7.20-7.35 (m, 6H), 7.40 (d, 2H), 7.55 (d, 2H), 7.65 (s, 1H), 7.75 (s,1H), 11.00 (s, 1H).

EXAMPLE 602-[(Aminocarbonyl)amino]-5-[4-{[N-(2-hydroxyethyl)-N-methylamino]methyl}phenyl]thiophene-3-carboxamide

2-[(Aminocarbonyl)amino]-5-[4-formylphenyl]thiophene-3-carboxamide (0.1g) was stirred in a mixture of DME (10 ml) and DMA(5 ml).2-(Methylamino)ethanol (0.13 g) was added, followed by trimethylorthoformate (5 ml) and acetic acid (0.5 ml). The reaction was stirredat 80° C. for 20 min, and then polymer-supported cyanoborohydride (0.45g) was added. The reaction was stirred at 80° C. for a further 2 h, andthen polymer-supported benzaldehyde (0.5 g) was added. The resins wereremoved by filtration and the filtrate was passed through a 5 g SCXcolumn, washing with methanol (25 ml). The product was eluted using 1Mmethanolic ammonia (45 ml) and this solution was evaporated to drynessunder reduced pressure. Purification by chromatography on silica,eluting with DCM/methanol (9:1), gave the product as an off-white solid(0.072 g).MS (ES) 349 (M+H)⁺.¹H NMR (DMSO-D6) 2.15 (s, 3H), 2.45 (m, 2H), 3.50 (m, 4H), 4.35 (t, 1H),6.90 (s, 2H), 7.25-7.40 (m, 3H), 7.50 (d, 2H), 7.70 (s, 1H), 7.75 (s,1H), 11.00 (s, 11).

EXAMPLE 612-[(Aminocarbonyl)amino]-5-[4-{([N-benzyl-N-(2-hydroxyethyl)]amino)methyl}phenyl]thiophene-3-carboxamide

The title compound was prepared using the general method of Example 60.

MS (ES) 425 (M+H)⁺.

¹H NMR (DMSO-D6) 2.45 (m, 2H), 3.50 (m, 2H), 3.60 (m, 4H), 4.40 (t, 1H),6.90 (s, 2H), 7.20-7.40 (m, 8H), 7.50 (d, 2H), 7.65 (s, 1H), 7.70 (s,1H), 11.00 (s, 1H).

EXAMPLE 622-[(Aminocarbonyl)amino]-5-[4-{(bis-[2-hydroxyethyl]amino)methyl}phenyl]thiophene-3-carboxamide

The title compound was prepared using the general method of Example 60.

MS (ES) 379 (M+H)⁺.

¹H NMR (DMSO-D6) 2.55 (1,4H), 3.45 (m, 4H), 3.60 (s, 2H), 4.35 (m, 2H),6.90 (s, 2H), 7.25 (s, 1H), 7.35 (d, 2H), 7.45 (d, 2H), 7.65 (s, 1H),7.75 (s, 1H), 11.00 (s, 1H).

Pharmacological Evaluation of Compounds

IKK-2 Filter Kinase Assay

Compounds were tested for inhibition of IKK-2 using a filter kinaseassay. The test compounds were dissolved to 10 mM in dimethylsulphoxide(DMSO). The compounds were then diluted 1 in 40 in kinase buffer (50 mMTris, pH 7.4 containing 0.1 mM EGTA, 0.1 mM sodium orthovanadate and0.1% β-mercaptoethanol). 1 in 3 serial dilutions were made from thissolution with 2.5% DMSO in kinase buffer. 20 μl of compound dilution wasadded to wells of a 96 well plate in duplicate. 20 μl 2.5% DMSO inkinase buffer instead of compound was added to control wells (0%inhibition). 20 μl 0.5 M EDTA was added instead of compound tobackground wells (100% inhibition).

10 μl of a mixture of magnesium acetate, unlabelled ATP, and³³P-labelled ATP was added to each well made such that the finalconcentration was 10 mM magnesium acetate, 1 μM ATP and 0.1 μCi ³³P ATP.20 μl of a mixture of IKK-2 (0.15 μg/well), 1-53 GST-IκB (0.5 μg/well)and bovine serum albumin (BSA) (8.5 μg/well) was added to each well tostart the reaction. The final reaction volume was 50 μl.

The kinase reactions were incubated at 21° C. for 80 minutes and thereaction stopped by precipitating the protein by the addition of anequal volume (50 μl) of 20% trichloroacetic acid (TCA). The precipitatewas allowed to form for 10 minutes and then filtered onto a GF/Cunifilter 96 well plate. Each filter was washed twice with approximately1 ml 2% TCA. The filter plate was dried at 30-40° C. for 60 minutes, 20μl scintillant was added to each well and the plate sealed andradioactivity counted on a Packard Topcount microplate scintillationcounter.

When tested in the above assay, the compounds of Examples 1 to 62 gaveIC₅₀ values of less than 10 μM indicating that they are expected to showuseful therapeutic activity.

IKK-1 Filter Kinase Assay

The selectivity of compounds was assessed by testing them for inhibitionof IKK-1 using a filter kinase assay. The assay conditions wereidentical to the IKK-2 filter kinase assay except that a mixture ofIKK-1 (0.25 μg/well) and 1-53 GST IκB (9 μg/well) was added to each wellto start the reaction.

Inhibition of LPS-induced TNFα production by PBMCs

The effect of test compounds on nuclear factor kappa B (NFκB) activationin cells was assessed by measuring inhibition of tumour necrosis factoralpha (TNFα) production by human peripheral blood mononuclear cells(PBMCs) stimulated by bacterial lipopolysaccharide (LPS).

Human blood (250 ml), anticoagulated with heparin, was collected fromhealthy volunteers. Aliquots of blood (25 ml) were layered on 20 mlLymphoprep (Nycomed) in 50 ml polypropylene centrifuge tubes. The tubeswere centrifuged (Sorval RT600B) at 2,500 rpm for 30 minutes. The cloudylayer containing PBMCs was collected with a fine tipped Pasteur pipette,transferred into 8 clean polypropylene centrifuge tubes (approximately10 ml per tube) and diluted to 50 ml with phosphate-buffered saline(PBS). These tubes were centrifuged at 2,000 rpm for 8 minutes. PBS (10ml) was added to each cell pellet and the cells were gentlyre-suspended. The cells were pooled in 4 centrifuge tubes, PBS was addedto each tube to make the volume up to 50 ml and the tubes werecentrifuged at 1,400 rpm for 8 minutes. The cell pellets were againre-suspended in 10 ml PBS, pooled in 2 centrige tubes, the volume madeup to 50 ml with PBS and the tubes centrifuged at 900 rpm for 10minutes.

The final cell pellets were gently re-suspended in 10 ml tissue culturemedium (RPMI containing 1% heat-inactivated human serum, L-glutamine andpenicillin and streptomycin), combined into 1 tube and the volume madeup to 30 ml with RPMI medium. The cells were counted and the cellsuspension was diluted to 2.6×10⁶ cells/ml.

Test compounds were dissolved in DMSO to 10 mM and diluted 1 in 250 (40μM) with RPMI medium. The compounds were then serially diluted 1 in 3with 0.4% DMSO in RPMI medium. Aliquots of test compound dilutions (50μl) were transferred to the wells of a 96-well plate. Control wellscontained 0.4% DMSO in RPMI instead of compound.

Aliquots of the cell suspension (100 μl) were added to each well and theplates incubated at 37° C. for 30 minutes. 50 μl of 40 μg/ml LPS (Sigma,LA130) was added to wells to stimulate TNFα production by the cells andthe plates were incubated overnight at 37° C. RPMI medium (50 μl) wasadded to negative control wells instead of LPS. The final incubationvolume was 200 μl.

Plates were centrifuged for 4 minutes at 1,200 rpm and supernatants wereremoved for measurement of TNFα concentration. Viability of theremaining cell pellet was measured using WST-1 reagent (BoehringerMannheim, 1044807). 100 μl RPMI medium containing 10 μl WST-1 reagentwas added to each well and the plates were incubated for 0.5 to 3 h. Theabsorbance at 450 nm was then measured using a 96-well platespectrophotometer.

TNFα in the supernatants (freshly harvested or stored frozen at −20° C.)were measured using an enzyme-linked immunosorbant assay (ELISA). TheELISA plate was prepared by coating the wells of a 96 well plate with asheep anti-human TNFα monoclonal antibody (100 μl of 1 μg/ml antibodydiluted in coating buffer; 0.5 M carbonate/bicarbonate buffer, pH 9.6containing 0.2 g/l sodium azide) and incubating overnight at 4° C. Blankwells were not coated. The wells were washed once with 0.1% BSA in PBScontaining 0.05% Tween (PBS/Tween) then incubated for 1 h at roomtemperature with 1% BSA in coating buffer (200 μl). The wells were thenwashed 3 times with 0.1% BSA in PBS/Tween.

The samples of supernatant from the PBMC incubation were diluted. 1 in 3with 1% BSA in PBS/Tween. 100 μl aliquots of these dilutions were addedto the ELISA plate. Other wells contained 100 μl TNFα standard (10, 3.3,1.1, 0.37, 0.12, 0.04, 0.014 and 0 ng/ml). The ELISA plate was incubatedat room temperature for 2 h before the wells were washed 3 times with0.1% BSA in PBS/Tween. A rabbit anti-human TNFα antibody (100 μl of a2.5 μg/ml solution) was added to each well and the plate incubated atroom temperature for 1.5 h. The wells were then washed 3 times with 0.1%BSA in PBS/Tween. Goat anti-rabbit IgG-horse radish peroxidase conjugate(ICN, 674371; 100 μl of a 1 in 10,000 dilution) was added to each welland the plate incubated at room temperature for 1.5 h. The wells werewashed 3 times with 0.1% BSA in PBS/Tween.

Peroxidase substrate was prepared by dissolving a 1 mg TMB tablet(Sigma, T-5525) in 100 μl DMSO (100 μl) and adding this and 36 μl UHPO(BDH, 30559; 1 g tablet dissolved in 25 ml distilled water) to 10 ml0.1M citrate/acetate buffer, pH6. 100 μl substrate was added to eachwell and the plate incubated in the dark at room temperature forapproximately 30 minutes. The reaction was stopped by adding 25 μl 2 Msulphuric acid to each well. The absorbance at 450 nm was measured in a96 well plater spectrophotometer.

Results

Activity in PBMC Inhibition of IKK-2 Assay Compound IC₅₀ (μM) IC₅₀ (μM)Example 1 0.0036 0.42 Example 5 0.013 0.12 Example 56 0.066 0.45 Example61 0.00056 0.34 Example 80, WO 01/58890 0.2 1.01 Example 82, WO 01/588900.26 1.15 Example 77, WO 01/58890 0.32 1.47

1. A compound of formula (I)

wherein R¹ represents H or CH₃; R² represents hydrogen, halogen, cyano,C1 to 2 alkyl, trifluoromethyl or C1 to 2 alkoxy; R³ and R⁴independently represent H or CH₃; A represents phenyl; and the group—CR³R⁴—X—R⁵ is bonded to ring A in the 4-position relative to thethiophene ring; X represents NR⁶; R⁵ represents H, C1 to 6 alkyl, C2 to6 alkenyl or C3 to 6 cycloalkyl; said alkyl group being optionallyfurther substituted by one or more groups selected independently fromCN, OH, C1 to 4 alkoxy, F, a C5 to 10 monocyclic or bicyclic aromaticring system and said ring system being optionally further substituted byone or more substituents selected independently from halogen, C1 to 2alkyl, C1 to 2 alkoxy or CF₃; or said alkyl being optionally furthersubstituted by a C5 to 6 cycloalkyl ring that optionally incorporates acarbonyl group and is optionally further substituted by OH; R⁶represents H or C1 to 6 alkyl; said alkyl group being optionally furthersubstituted by CN, OH, C1 to 4 alkoxy or one or more fluoro atoms; n andm independently represent an integer 0, 1 or 2; R⁷ and R⁸ independentlyrepresent H or C1 to 2 alkyl; and pharmaceutically acceptable saltsthereof.
 2. A compound of formula (I), according to claim 1, wherein R¹represents H.
 3. A compound of formula (I), according to claim 1, inwhich A represents optionally substituted phenyl.
 4. A compound offormula (I), according to claim 1, in which R³ and R⁴ each represent H.5. A process for the preparation of a compound of formula (I), accordingto claim 1, which comprises: (a) reaction of a compound of formula (II):

wherein A, R¹, R², R³, R⁴, R⁵ and X are as defined in claim 1 with anisocyanate; or (b) reaction of a compound of formula (III)

wherein A, R², R³, R⁴, R⁵ and X are as defined in claim 1, with acompound of formula (IV)

wherein R¹ is as defined in claim 1 and LG represents a leaving group;or (c) reaction of a compound of formula (V)

wherein A, R², R³, R⁴, R⁵ and X are as defined in claim 1 and LGrepresents a leaving group, with a compound of formula (VI)

wherein R¹ is as defined in claim 1; or (d) reaction of a compound offormula (VII)

wherein A, R¹, R², R³ and R⁴ are as defined in claim 1, and LGrepresents a leaving group, with an amine of formula R⁵R⁶NH, wherein R⁵and R⁶ are as defined in claim 1; or (e) reaction of a compound offormula (VIII)

wherein A, R¹, R² and R³ are as defined in claim 1, with an amine offormula R⁵R⁶NH wherein R⁵ and R⁶ are as defined in claim 1, underreductive amination conditions; or (f) reaction of a compound of formula(IX)

wherein R¹, R², R³, R⁴, R⁵ and A are as defined in claim 1, with analdehyde or ketone under reductive amination conditions; and wherenecessary converting the resultant compound of formula (I), or anothersalt thereof, into a pharmaceutically acceptable salt thereof; orconverting the resultant compound of formula (I) into a further compoundof formula (I); and where desired converting the resultant compound offormula (I) into an optical isomer thereof.
 6. A pharmaceuticalcomposition comprising a compound of formula (I), or a pharmaceuticallyacceptable salt thereof; as claimed in claim 1 in association with apharmaceutically acceptable adjuvant, diluent or carrier.
 7. Apharmaceutical composition adapted for administration by inhalation orinsufflation comprising a compound of formula (I), or a pharmaceuticallyacceptable salt thereof, as claimed in claim 1 in association with apharmaceutically acceptable adjuvant, diluent or carrier.
 8. A processfor the preparation of a pharmaceutical composition which comprisesmixing a compound of formula (I), or a pharmaceutically acceptable saltthereof, as claimed claim 1 with a pharmaceutically acceptable adjuvant,diluent or carrier.
 9. A method for the treatment or prophylaxis ofinflammatory disease selected from the group consisting of asthma,rheumatoid arthritis, psoriasis, inflammatory bowel disease, multiplesclerosis, chronic obstructive pulmonary disease, bone resorptivedisease, osteoarthritis, and diabetes/glycemic control, comprisingadministering to a person suffering from or at risk of said disease atherapeutically effective amount of a compound of formula (I), or apharmaceutically acceptable salt thereof, as claimed in claim
 1. 10. Themethod as claimed in claim 9 wherein the disease is rheumatoidarthritis.
 11. The method as claimed in claim 9 wherein the disease ischronic obstructive pulmonary disease.
 12. A method of treating, orreducing the risk of cancer wherein the cancer is selected from thegroup consisting of colorectal cancer, breast cancer, gastric cancer,Hodgkin's lymphoma, head and neck squamous cell carcinoma, ChronicMyelogenous Leukemia, and Acute Lymphoblastic Leukemia, which comprisesadministering to a person suffering from or at risk of said disease orcondition a therapeutically effective amount of a compound of formula(I), or a pharmaceutically acceptable salt thereof, as claimed inclaim
 1. 13. A compound of formula (I), according to claim 2, in which Arepresents optionally substituted phenyl.
 14. A compound of formula (I),according to claim 2, in which R³ and R⁴ each represent H.
 15. Acompound of formula (I), according to claim 3, in which R³ and R⁴ eachrepresent H.
 16. A compound of formula (I), according to claim 13, inwhich R³ and R⁴ each represent H.